Centro Andaluz de Biología Molecular (CABIMER)

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  • Publication
    Bound galloylated compounds in persimmon upcycled dietary fiber modulate microbial strains associated to human health after in vitro digestion
    (Elsevier, 2021-12-28) Moreno-Chamba, Bryan; Salazar-Bermeo, Julio; Concepcion Martinez-Madrid, Maria; Lizama, Victoria; Martin-Bermudo, Francisco; Berna, Genoveva; Neacsu, Madalina; Saura, Domingo; Marti, Nuria; Valero, Manuel; [Moreno-Chamba, Bryan] Univ Miguel Hernandez Elche UMH, Inst Invest Desarrollo & Innovat Biotecnol Sanita, Edificio Torregaitan,Ave Univ S-N, Alicante 03202, Spain; [Salazar-Bermeo, Julio] Univ Miguel Hernandez Elche UMH, Inst Invest Desarrollo & Innovat Biotecnol Sanita, Edificio Torregaitan,Ave Univ S-N, Alicante 03202, Spain; [Saura, Domingo] Univ Miguel Hernandez Elche UMH, Inst Invest Desarrollo & Innovat Biotecnol Sanita, Edificio Torregaitan,Ave Univ S-N, Alicante 03202, Spain; [Marti, Nuria] Univ Miguel Hernandez Elche UMH, Inst Invest Desarrollo & Innovat Biotecnol Sanita, Edificio Torregaitan,Ave Univ S-N, Alicante 03202, Spain; [Valero, Manuel] Univ Miguel Hernandez Elche UMH, Inst Invest Desarrollo & Innovat Biotecnol Sanita, Edificio Torregaitan,Ave Univ S-N, Alicante 03202, Spain; [Concepcion Martinez-Madrid, Maria] Univ Miguel Hernandez Elche UMH, Dept Agroquim & Medio Ambiente, Campus Orihuela,Carretera Beniel Km 3-2, Alicante 03312, Spain; [Lizama, Victoria] Univ Politecn Valencia, Inst Ingn Alimentos Desarrollo, Ave Fausto Elio S-N,Edificio 8E,Acceso F Planta 0, Valencia 46022, Spain; [Martin-Bermudo, Francisco] Univ Seville, Univ Pablo Olavide, Ctr Andaluz Biol Mol & Med Regenerat CABIMER, Consejo Super Invest Cient CSIC, Seville 41092, Spain; [Berna, Genoveva] Univ Seville, Univ Pablo Olavide, Ctr Andaluz Biol Mol & Med Regenerat CABIMER, Consejo Super Invest Cient CSIC, Seville 41092, Spain; [Neacsu, Madalina] Univ Aberdeen, Rowett Res Inst, Ashgrove Rd W, Aberdeen AB25 2ZD, Scotland; Ministry of Science Innovation and Universities
    Persimmon byproduct upcycling was performed by solvent-assisted extraction (SAE) to obtain dietary fiber (DF). The effect of SAE on DF modulation was studied on specific beneficial and pathogenic strains before and after the in vitro digestion process. Overall, digested DF samples extracted using acetone as a solvent (dCET) showed higher (p
  • Publication
    SENP7 overexpression protects cancer cells from oxygen and glucose deprivation and associates with poor prognosis in colon cancer.
    (2022-04-01) Gallardo-Chamizo, Francisco; Lara-Ureña, Nieves; Correa-Vázquez, Juan F; Reyes, José C; Gauthier, Benoit R; García-Domínguez, Mario
  • Publication
    Metabolic-associated fatty liver disease: From simple steatosis toward liver cirrhosis and potential complications. Proceedings of the Third Translational Hepatology Meeting, organized by the Spanish Association for the Study of the Liver (AEEH).
    (2022-03-03) Gallego-Durán, Rocío; Albillos, Agustín; Ampuero, Javier; Arechederra, María; Bañares, Rafael; Blas-García, Ana; Berná, Genoveva; Caparrós, Esther; Delgado, Teresa C; Falcón-Pérez, Juan Manuel; Francés, Rubén; Fernández-Barrena, Maite G; Graupera, Isabel; Iruzubieta, Paula; Nevzorova, Yulia A; Nogueiras, Rubén; Macías, Rocío I R; Martín, Franz; Sabio, Guadalupe; Soriano, Germán; Vaquero, Javier; Cubero, Francisco Javier; Gracia-Sancho, Jordi
    This is a meeting report of the 3rd Translational Hepatology Meeting held in Alicante, Spain, in October 2021. The meeting, which was organized by the Spanish Association for the Study of the Liver (AEEH), provided an update on the recent advances in the field of basic and translational hepatology, with a particular focus on the molecular and cellular mechanisms and therapeutic targets involved in metabolic-associated fatty liver disease (MAFLD), metabolic-associated steatohepatitis (MASH), cirrhosis and end-stage hepatocellular carcinoma (HCC).
  • Publication
    Data of transcriptional effects of the merbarone-mediated inhibition of TOP2.
    (2022-08-01) Delgado-Chaves, Fernando M; Martínez-García, Pedro Manuel; Herrero-Ruiz, Andrés; Gómez-Vela, Francisco; Divina, Federico; Jimeno-González, Silvia; Cortés-Ledesma, Felipe
    Type II DNA topoisomerases relax topological stress by transiently gating DNA passage in a controlled cut-and-reseal mechanism that affects both DNA strands. Therefore, they are essential to overcome topological problems associated with DNA metabolism. Their aberrant activity results in the generation of DNA double-strand breaks, which can seriously compromise cell survival and genome integrity. Here, we profile the transcriptome of human-telomerase-immortalized retinal pigment epithelial 1 (RPE-1) cells when treated with merbarone, a drug that catalytically inhibits type II DNA topoisomerases. We performed RNA-Seq after 4 and 8 h of merbarone treatment and compared transcriptional profiles versus untreated samples. We report raw sequencing data together with lists of gene counts and differentially expressed genes.
  • Publication
    Topoisomerase 1-dependent R-loop deficiency drives accelerated replication and genomic instability.
    (2022) Sarni, Dan; Barroso, Sonia; Shtrikman, Alon; Irony-Tur Sinai, Michal; Oren, Yifat S; Aguilera, Andrés; Kerem, Batsheva
    DNA replication is a complex process tightly regulated to ensure faithful genome duplication, and its perturbation leads to DNA damage and genomic instability. Replication stress is commonly associated with slow and stalled replication forks. Recently, accelerated replication has emerged as a non-canonical form of replication stress. However, the molecular basis underlying fork acceleration is largely unknown. Here, we show that mutated HRAS activation leads to increased topoisomerase 1 (TOP1) expression, causing aberrant replication fork acceleration and DNA damage by decreasing RNA-DNA hybrids or R-loops. In these cells, restoration of TOP1 expression or mild replication inhibition rescues the perturbed replication and reduces DNA damage. Furthermore, TOP1 or RNaseH1 overexpression induces accelerated replication and DNA damage, highlighting the importance of TOP1 equilibrium in regulating R-loop homeostasis to ensure faithful DNA replication and genome integrity. Altogether, our results dissect a mechanism of oncogene-induced DNA damage by aberrant replication fork acceleration.
  • Publication
    MutSβ regulates G4-associated telomeric R-loops to maintain telomere integrity in ALT cancer cells.
    (2022) Sakellariou, Despoina; Bak, Sara Thornby; Isik, Esin; Barroso, Sonia I; Porro, Antonio; Aguilera, Andrés; Bartek, Jiri; Janscak, Pavel; Peña-Diaz, Javier
    Up to 15% of human cancers maintain their telomeres through a telomerase-independent mechanism, termed "alternative lengthening of telomeres" (ALT) that relies on homologous recombination between telomeric sequences. Emerging evidence suggests that the recombinogenic nature of ALT telomeres results from the formation of RNA:DNA hybrids (R-loops) between telomeric DNA and the long-noncoding telomeric repeat-containing RNA (TERRA). Here, we show that the mismatch repair protein MutSβ, a heterodimer of MSH2 and MSH3 subunits, is enriched at telomeres in ALT cancer cells, where it prevents the accumulation of telomeric G-quadruplex (G4) structures and R-loops. Cells depleted of MSH3 display increased incidence of R-loop-dependent telomere fragility and accumulation of telomeric C-circles. We also demonstrate that purified MutSβ recognizes and destabilizes G4 structures in vitro. These data suggest that MutSβ destabilizes G4 structures in ALT telomeres to regulate TERRA R-loops, which is a prerequisite for maintenance of telomere integrity during ALT.
  • Publication
    Abnormal cannabidiol ameliorates inflammation preserving pancreatic beta cells in mouse models of experimental type 1 diabetes and beta cell damage.
    (2021-12-03) González-Mariscal, Isabel; Pozo-Morales, Macarena; Romero-Zerbo, Silvana Y; Espinosa-Jimenez, Vanesa; Escamilla-Sánchez, Alejandro; Sánchez-Salido, Lourdes; Cobo-Vuilleumier, Nadia; Gauthier, Benoit R; Bermúdez-Silva, Francisco J
    The atypical cannabinoid Abn-CBD improves the inflammatory status in preclinical models of several pathologies, including autoimmune diseases. However, its potential for modulating inflammation in autoimmune type 1 diabetes (T1D) is unknown. Herein we investigate whether Abn-CBD can modulate the inflammatory response during T1D onset using a mouse model of T1D (non-obese diabetic- (NOD)-mice) and of beta cell damage (streptozotocin (STZ)-injected mice). Six-week-old female NOD mice were treated with Abn-CBD (0.1-1 mg/kg) or vehicle during 12 weeks and then euthanized. Eight-to-ten-week-old male C57Bl6/J mice were pre-treated with Abn-CBD (1 mg/kg of body weight) or vehicle for 1 week, following STZ challenge, and euthanized 1 week later. Blood, pancreas, pancreatic lymph nodes (PLNs) and T cells were collected and processed for analysis. Glycemia was also monitored. In NOD mice, treatment with Abn-CBD significantly reduced the severity of insulitis and reduced the pro-inflammatory profile of CD4+ T cells compared to vehicle. Concomitantly, Abn-CBD significantly reduced islet cell apoptosis and improved glucose tolerance. In STZ-injected mice, Abn-CBD decreased circulating proinflammatory cytokines and ameliorated islet inflammation reducing intra-islet phospho-NF-κB and TXNIP. Abn-CBD significantly reduced 2 folds intra-islet CD8+ T cells and reduced Th1/non-Th1 ratio in PLNs of STZ-injected mice. Islet cell apoptosis and intra-islet fibrosis were also significantly reduced in Abn-CBD pre-treated mice compared to vehicle. Altogether, Abn-CBD reduces circulating and intra-islet inflammation, preserving islets, thus delaying the progression of insulitis. Hence, Abn-CBD and related compounds emerge as new candidates to develop pharmacological strategies to treat the early stages of T1D.
  • Publication
    Telomeric C-circles localize at nuclear pore complexes in Saccharomyces cerevisiae.
    (2022-02-11) Aguilera, Paula; Dubarry, Marion; Hardy, Julien; Lisby, Michael; Simon, Marie-Noëlle; Géli, Vincent
    As in human cells, yeast telomeres can be maintained in cells lacking telomerase activity by recombination-based mechanisms known as ALT (Alternative Lengthening of Telomeres). A hallmark of ALT human cancer cells are extrachromosomal telomeric DNA elements called C-circles, whose origin and function have remained unclear. Here, we show that extrachromosomal telomeric C-circles in yeast can be detected shortly after senescence crisis and concomitantly with the production of survivors arising from "type II" recombination events. We uncover that C-circles bind to the nuclear pore complex (NPC) and to the SAGA-TREX2 complex, similar to other non-centromeric episomal DNA. Disrupting the integrity of the SAGA/TREX2 complex affects both C-circle binding to NPCs and type II telomere recombination, suggesting that NPC tethering of C-circles facilitates formation and/or propagation of the long telomere repeats characteristic of type II survivors. Furthermore, we find that disruption of the nuclear diffusion barrier impairs type II recombination. These results support a model in which concentration of C-circles at NPCs benefits type II telomere recombination, highlighting the importance of spatial coordination in ALT-type mechanisms of telomere maintenance.
  • Publication
    Manganese is a physiologically relevant TORC1 activator in yeast and mammals.
    (2022-07-29) Nicastro, Raffaele; Gaillard, Hélène; Zarzuela, Laura; Péli-Gulli, Marie-Pierre; Fernández-García, Elisabet; Tomé, Mercedes; García-Rodríguez, Néstor; Durán, Raúl V; De Virgilio, Claudio; Wellinger, Ralf Erik
    The essential biometal manganese (Mn) serves as a cofactor for several enzymes that are crucial for the prevention of human diseases. Whether intracellular Mn levels may be sensed and modulate intracellular signaling events has so far remained largely unexplored. The highly conserved target of rapamycin complex 1 (TORC1, mTORC1 in mammals) protein kinase requires divalent metal cofactors such as magnesium (Mg2+) to phosphorylate effectors as part of a homeostatic process that coordinates cell growth and metabolism with nutrient and/or growth factor availability. Here, our genetic approaches reveal that TORC1 activity is stimulated in vivo by elevated cytoplasmic Mn levels, which can be induced by loss of the Golgi-resident Mn2+ transporter Pmr1 and which depend on the natural resistance-associated macrophage protein (NRAMP) metal ion transporters Smf1 and Smf2. Accordingly, genetic interventions that increase cytoplasmic Mn2+ levels antagonize the effects of rapamycin in triggering autophagy, mitophagy, and Rtg1-Rtg3-dependent mitochondrion-to-nucleus retrograde signaling. Surprisingly, our in vitro protein kinase assays uncovered that Mn2+ activates TORC1 substantially better than Mg2+, which is primarily due to its ability to lower the Km for ATP, thereby allowing more efficient ATP coordination in the catalytic cleft of TORC1. These findings, therefore, provide both a mechanism to explain our genetic observations in yeast and a rationale for how fluctuations in trace amounts of Mn can become physiologically relevant. Supporting this notion, TORC1 is also wired to feedback control mechanisms that impinge on Smf1 and Smf2. Finally, we also show that Mn2+-mediated control of TORC1 is evolutionarily conserved in mammals, which may prove relevant for our understanding of the role of Mn in human diseases.
  • Publication
    Heterogeneity of In Vitro Expanded Mesenchymal Stromal Cells and Strategies to Improve Their Therapeutic Actions.
    (2022-05-23) Olmedo-Moreno, Laura; Aguilera, Yolanda; Baliña-Sánchez, Carmen; Martín-Montalvo, Alejandro; Capilla-González, Vivian
    Beneficial properties of mesenchymal stromal cells (MSCs) have prompted their use in preclinical and clinical research. Accumulating evidence has been provided for the therapeutic effects of MSCs in several pathologies, including neurodegenerative diseases, myocardial infarction, skin problems, liver disorders and cancer, among others. Although MSCs are found in multiple tissues, the number of MSCs is low, making in vitro expansion a required step before MSC application. However, culture-expanded MSCs exhibit notable differences in terms of cell morphology, physiology and function, which decisively contribute to MSC heterogeneity. The changes induced in MSCs during in vitro expansion may account for the variability in the results obtained in different MSC-based therapy studies, including those using MSCs as living drug delivery systems. This review dissects the different changes that occur in culture-expanded MSCs and how these modifications alter their therapeutic properties after transplantation. Furthermore, we discuss the current strategies developed to improve the beneficial effects of MSCs for successful clinical implementation, as well as potential therapeutic alternatives.
  • Publication
    TGF-β-Upregulated Lnc-Nr6a1 Acts as a Reservoir of miR-181 and Mediates Assembly of a Glycolytic Complex.
    (2022-09-15) Polo-Generelo, Salvador; Torres, Belén; Guerrero-Martínez, José A; Camafeita, Emilio; Vázquez, Jesús; Reyes, José C; Pintor-Toro, José A
    Long non-coding RNAs (lncRNAs) have emerged as key regulators in a wide range of biological processes. Here, we identified a mouse miRNA-host gene lncRNA (lnc-Nr6a1) upregulated early during epithelial-to-mesenchymal transition (EMT). We show that when lncRNA is processed, it gives rise to two abundant polyadenylated isoforms, lnc-Nr6a1-1 and lnc-Nr6a1-2, and a longer non-polyadenylated microprocessor-driven lnc-pri-miRNA containing clustered pre-miR-181a2 and pre-miR-181b2 hairpins. Ectopic expression of the lnc-Nr6a1-1 or lnc-Nr6a1-2 isoform enhanced cell migration and the invasive capacity of the cells, whereas the expression of the isoforms and miR-181a2 and miR-181b2 conferred anoikis resistance. Lnc-Nr6a1 gene deletion resulted in cells with lower adhesion capacity and reduced glycolytic metabolism, which are restored by lnc-Nr6a1-1 isoform expression. We performed identification of direct RNA interacting proteins (iDRIP) to identify proteins interacting directly with the lnc-Nr6a1-1 isoform. We defined a network of interacting proteins, including glycolytic enzymes, desmosome proteins and chaperone proteins; and we demonstrated that the lnc-Nr6a1-1 isoform directly binds and acts as a scaffold molecule for the assembly of ENO1, ALDOA, GAPDH, and PKM glycolytic enzymes, along with LDHA, supporting substrate channeling for efficient glycolysis. Our results unveil a role of Lnc-Nr6a1 as a multifunctional lncRNA acting as a backbone for multiprotein complex formation and primary microRNAs.
  • Publication
    Adaptive Response of Saccharomyces Hosts to Totiviridae L-A dsRNA Viruses Is Achieved through Intrinsically Balanced Action of Targeted Transcription Factors.
    (2022-04-09) Ravoitytė, Bazilė; Lukša, Juliana; Wellinger, Ralf Erik; Serva, Saulius; Servienė, Elena
    Totiviridae L-A virus is a widespread yeast dsRNA virus. The persistence of the L-A virus alone appears to be symptomless, but the concomitant presence of a satellite M virus provides a killer trait for the host cell. The presence of L-A dsRNA is common in laboratory, industrial, and wild yeasts, but little is known about the impact of the L-A virus on the host's gene expression. In this work, based on high-throughput RNA sequencing data analysis, the impact of the L-A virus on whole-genome expression in three different Saccharomyces paradoxus and S. cerevisiae host strains was analyzed. In the presence of the L-A virus, moderate alterations in gene expression were detected, with the least impact on respiration-deficient cells. Remarkably, the transcriptional adaptation of essential genes was limited to genes involved in ribosome biogenesis. Transcriptional responses to L-A maintenance were, nevertheless, similar to those induced upon stress or nutrient availability. Based on these data, we further dissected yeast transcriptional regulators that, in turn, modulate the cellular L-A dsRNA levels. Our findings point to totivirus-driven fine-tuning of the transcriptional landscape in yeasts and uncover signaling pathways employed by dsRNA viruses to establish the stable, yet allegedly profitless, viral infection of fungi.
  • Publication
    Mesoporous Silica-Based Nanoparticles as Non-Viral Gene Delivery Platform for Treating Retinitis Pigmentosa.
    (2022-04-13) Valdés-Sánchez, Lourdes; Borrego-González, Sara; Montero-Sánchez, Adoración; Massalini, Simone; de la Cerda, Berta; Díaz-Cuenca, Aránzazu; Díaz-Corrales, Francisco J
    Gene therapy is a therapeutic possibility for retinitis pigmentosa (RP), in which therapeutic transgenes are currently delivered to the retina by adeno-associated viral vectors (AAVs). Although their safety and efficacy have been demonstrated in both clinical and preclinical settings, AAVs present some technical handicaps, such as limited cargo capacity and possible immunogenicity in repetitive doses. The development of alternative, non-viral delivery platforms like nanoparticles is of great interest to extend the application of gene therapy for RP. Amino-functionalized mesoporous silica-based nanoparticles (N-MSiNPs) were synthesized, physico-chemically characterized, and evaluated as gene delivery systems for human cells in vitro and for retinal cells in vivo. Transgene expression was evaluated by WB and immunofluorescence. The safety evaluation of mice subjected to subretinal injection was assessed by ophthalmological tests (electroretinogram, funduscopy, tomography, and optokinetic test). N-MSiNPs delivered transgenes to human cells in vitro and to retinal cells in vivo. No adverse effects were detected for the integrity of the retinal tissue or the visual function of treated eyes. N-MSiNPs were able to deliver a therapeutic transgene candidate for RP, PRPF31, both in vitro and in vivo. N-MSiNPs are safe for retinal delivery and thus a potential alternative to viral vectors.
  • Publication
    Restoring TRAILR2/DR5-Mediated Activation of Apoptosis upon Endoplasmic Reticulum Stress as a Therapeutic Strategy in Cancer.
    (2022-08-12) Mora-Molina, Rocío; López-Rivas, Abelardo
    The uncontrolled proliferation of malignant cells in growing tumors results in the generation of different stressors in the tumor microenvironment, such as nutrient shortage, hypoxia and acidosis, among others, that disrupt endoplasmic reticulum (ER) homeostasis and may lead to ER stress. As a response to ER stress, both normal and tumor cells launch a set of signaling pathways known as the unfolded protein response (UPR) to restore ER proteostasis and maintain cell viability and function. However, under sustained ER stress, an apoptotic cell death process can be induced and this has been the subject of different review articles, although the role of the TRAIL-R2/DR5-activated extrinsic pathway of apoptosis has not yet been thoroughly summarized. In this Review, we provide an updated overview of the molecular mechanisms regulating cell fate decisions in tumor cells undergoing ER stress and discuss the role of the tumor necrosis factor (TNF)-related apoptosis-inducing ligand receptor 2 (TRAIL-R2/DR5) in the final outcome of UPR signaling. Particularly, we focus on the mechanisms controlling cellular FLICE-like inhibitory protein (FLIP) levels in tumor cells undergoing ER stress, which may represent a potential target for therapeutic intervention in cancer.
  • Publication
    Cancer-Associated Dysregulation of Sumo Regulators: Proteases and Ligases.
    (2022-07-20) Lara-Ureña, Nieves; Jafari, Vahid; García-Domínguez, Mario
    SUMOylation is a post-translational modification that has emerged in recent decades as a mechanism involved in controlling diverse physiological processes and that is essential in vertebrates. The SUMO pathway is regulated by several enzymes, proteases and ligases being the main actors involved in the control of sumoylation of specific targets. Dysregulation of the expression, localization and function of these enzymes produces physiological changes that can lead to the appearance of different types of cancer, depending on the enzymes and target proteins involved. Among the most studied proteases and ligases, those of the SENP and PIAS families stand out, respectively. While the proteases involved in this pathway have specific SUMO activity, the ligases may have additional functions unrelated to sumoylation, which makes it more difficult to study their SUMO-associated role in cancer process. In this review we update the knowledge and advances in relation to the impact of dysregulation of SUMO proteases and ligases in cancer initiation and progression.
  • Publication
    Laparoscopic Sleeve Gastrectomy in Patients with Severe Obesity Restores Adaptive Responses Leading to Nonalcoholic Steatohepatitis.
    (2022-07-15) Cabré, Noemí; Luciano-Mateo, Fedra; Chapski, Douglas J; Baiges-Gaya, Gerard; Fernández-Arroyo, Salvador; Hernández-Aguilera, Anna; Castañé, Helena; Rodríguez-Tomàs, Elisabet; París, Marta; Sabench, Fàtima; Castillo, Daniel Del; Del Bas, Josep M; Tomé, Mercedes; Bodineau, Clément; Sola-García, Alejandro; López-Miranda, José; Martín-Montalvo, Alejandro; Durán, Raúl V; Vondriska, Thomas M; Rosa-Garrido, Manuel; Camps, Jordi; Menéndez, Javier A; Joven, Jorge
    The surgically induced remission of liver disease represents a model to investigate the signalling processes that trigger the development of nonalcoholic steatohepatitis with the aim of identifying novel therapeutic targets. We recruited patients with severe obesity with or without nonalcoholic steatohepatitis and obtained liver and plasma samples before and after laparoscopic sleeve gastrectomy for immunoblotting, immunocytochemical, metabolomic, transcriptomic and epigenetic analyses. Functional studies were performed in HepG2 cells and primary hepatocytes. Surgery was associated with a decrease in the inflammatory response and revealed the role of mitogen-activated protein kinases. Nonalcoholic steatohepatitis was associated with an increased glutaminolysis-induced production of α-ketoglutarate and the hyperactivation of mammalian target of rapamycin complex 1. These changes were crucial for adenosine monophosphate-activated protein kinase/mammalian target of rapamycin-driven pathways that modulated hepatocyte survival by coordinating apoptosis and autophagy and affected methylation-related epigenomic remodelling enzymes. Hepatic transcriptome signatures and differentially methylated genomic regions distinguished patients with and without steatohepatitis. Our results suggest that the increased glutaminolysis-induced α-ketoglutarate production and the mammalian target of rapamycin complex 1 dysregulation play a crucial role in the inefficient adaptive responses leading to steatohepatitis in obesity.
  • Publication
    Human Omental Mesothelial Cells Impart an Immunomodulatory Landscape Impeding B- and T-Cell Activation.
    (2022-05-25) Gauthier, Benoit R; Rubio-Contreras, Diana; Gómez-Rosado, Juan Carlos; Capitán-Morales, Luis Cristobal; Hmadcha, Abdelkrim; Soria, Bernat; Lachaud, Christian Claude
    Mesothelial cells form the mesothelium, a simple epithelium lining the walls of serous cavities and the surface of visceral organs. Although mesothelial cells are phenotypically well characterized, their immunoregulatory properties remain largely unknown, with only two studies reporting their capacity to inhibit T cells through TGF-β and their consumption of L-arginine by arginase-1. Whether human mesothelial cells can suppress other immune cells and possess additional leukosuppressive mechanisms, remain to be addressed to better delineate their therapeutic potential for cell therapy. Herein, we generated secretomes from omental mesothelial cells (OMC) and assess their capacity to inhibit lymphocytes proliferation, suppress activated T and B cells, as well as to modify macrophage activation markers. The secretome from mesenchymal stromal cells (MSC) served as a control of immuno-suppression. Although OMC and MSC were phenotypically divergent, their cytokine secretion patterns as well as expression of inflammatory and immunomodulary genes were similar. As such, OMC- and MSC-derived secretomes (OMC-S and MSC-S) both polarized RAW 264.7 macrophages towards a M2-like anti-inflammatory phenotype and suppressed mouse and human lymphocytes proliferation. OMC-S displayed a strong ability to suppress mouse- and human-activated CD19+/CD25+ B cells as compared to MSC-S. The lymphosuppressive activity of the OMC-S could be significantly counteracted either by SB-431542, an inhibitor of TGFβ and activin signaling pathways, or with a monoclonal antibody against the TGFβ1, β2, and β3 isoforms. A strong blockade of the OMC-S-mediated lymphosuppressive activity was achieved using L-NMMA, a specific inhibitor of nitric oxide synthase (NOS). Taken together, our results suggest that OMC are potent immunomodulators.
  • Publication
    Titanium Surface Characteristics Induce the Specific Reprogramming of Toll-like Receptor Signaling in Macrophages.
    (2022-04-13) González-Sánchez, Zaira; Areal-Quecuty, Victoria; Jimenez-Guerra, Alvaro; Cabanillas-Balsera, Daniel; Gil, Francisco Javier; Velasco-Ortega, Eugenio; Pozo, David
    Most of the research on titanium-based dental implants (Ti-discs) is focused on how they are able to stimulate the formation of new tissue and/or cytotoxic studies, with very scarce data on their effects on functional responses by immunocompetent cells. In particular, the link between the rewiring of innate immune responses and surface biomaterials properties is poorly understood. To address this, we characterize the functional response of macrophage cultures to four different dental titanium surfaces (MA: mechanical abrasion; SB + AE: sandblasting plus etching; SB: sandblasting; AE: acid etching). We use different Toll-like receptor (TLR) ligands towards cell surface receptors (bacterial lipopolysaccharide LPS for TLR4; imiquimod for TLR7; synthetic bacterial triacylated lipoprotein for TLR2/TLR1) and endosomal membrane receptor (poly I:C for TLR3) to simulate bacterial (cell wall bacterial components) or viral infections (dsRNA and ssRNA). The extracellular and total LDH levels indicate that exposure to the different Ti-surfaces is not cytotoxic for macrophages under resting or TLR-stimulated conditions, although there is a tendency towards an impairment in macrophage proliferation, viability or adhesion under TLR4, TLR3 and TLR2/1 stimulations in SB discs cultures. The secreted IL-6 and IL-10 levels are not modified upon resting macrophage exposure to the Ti-surfaces studied as well as steady state levels of iNos or ArgI mRNA. However, macrophage exposure to MA Ti-surface do display an enhanced immune response to TLR4, TLR7 or TLR2/1 compared to other Ti-surfaces in terms of soluble immune mediators secreted and M1/M2 gene expression profiling. This change of characteristics in cellular phenotype might be related to changes in cellular morphology. Remarkably, the gene expression of Tlr3 is the only TLR that is differentially affected by distinct Ti-surface exposure. These results highlight the relevance of patterned substrates in dental implants to achieve a smart manipulation of the immune responses in the context of personalized medicine, cell-based therapies, preferential lineage commitment of precursor cells or control of tissue architecture in oral biology.
  • Publication
    Pdx1 Is Transcriptionally Regulated by EGR-1 during Nitric Oxide-Induced Endoderm Differentiation of Mouse Embryonic Stem Cells.
    (2022-04-01) Salguero-Aranda, Carmen; Beltran-Povea, Amparo; Postigo-Corrales, Fátima; Hitos, Ana Belén; Díaz, Irene; Caballano-Infantes, Estefanía; Fraga, Mario F; Hmadcha, Abdelkrim; Martín, Franz; Soria, Bernat; Tapia-Limonchi, Rafael; Bedoya, Francisco J; Tejedo, Juan R; Cahuana, Gladys M
    The transcription factor, early growth response-1 (EGR-1), is involved in the regulation of cell differentiation, proliferation, and apoptosis in response to different stimuli. EGR-1 is described to be involved in pancreatic endoderm differentiation, but the regulatory mechanisms controlling its action are not fully elucidated. Our previous investigation reported that exposure of mouse embryonic stem cells (mESCs) to the chemical nitric oxide (NO) donor diethylenetriamine nitric oxide adduct (DETA-NO) induces the expression of early differentiation genes such as pancreatic and duodenal homeobox 1 (Pdx1). We have also evidenced that Pdx1 expression is associated with the release of polycomb repressive complex 2 (PRC2) and P300 from the Pdx1 promoter; these events were accompanied by epigenetic changes to histones and site-specific changes in the DNA methylation. Here, we investigate the role of EGR-1 on Pdx1 regulation in mESCs. This study reveals that EGR-1 plays a negative role in Pdx1 expression and shows that the binding capacity of EGR-1 to the Pdx1 promoter depends on the methylation level of its DNA binding site and its acetylation state. These results suggest that targeting EGR-1 at early differentiation stages might be relevant for directing pluripotent cells into Pdx1-dependent cell lineages.