Articles publicats en revistes (Biomedicina)

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Preclinical study of microphthalmia-associated transcription factor inhibitor ML329 in gastrointestinal stromal tumor growth
(Elsevier, 2025-04-14) Guerrero, Mario; Proaño Pérez, Elisabeth; Serrano Candelas, Eva; García Valverde, Alfonso; Carrillo Rodríguez, Berenice; Rosell, Jordi; Serrano, César; Martín Andorrà, Margarita
Gastrointestinal stromal tumors (GISTs) comprise about 80% of mesenchymal neoplasms in the gastrointestinal tract. Although imatinib mesylate is the preferred treatment, the development of drug resistance highlights the need for novel therapeutic strategies. Recently, we have identified the microphthalmia-associated transcription factor (MITF) as a critical player in pro-survival signaling and tumor growth. This study investigates the effects of MITF inhibition using ML329, an MITF pathway inhibitor, on GIST cell viability in vitro and in NMRI-nu/nu mouse xenograft models. ML329 suppresses growth in imatinib-sensitive (GIST-T1) and -resistant (GIST 430/654) cell lines, impairs MITF targets such as BCL2 and CDK2, and induces S-G2/M cell-cycle arrest. In vivo, ML329 is well tolerated and significantly reduces tumor growth in established imatinib-sensitive and -resistant GIST models. These findings underscore the importance of MITF in GIST growth and support its inhibition as a promising therapeutic approach.
Liver X receptors and inflammatory-induced C/EBPβ selectively cooperate to control CD38 transcription
(Karger, 2024-12-19) Glaría Percaz, Estibaliz; Rodríguez Martínez, Pol; Font Díaz, Joan; Rosa, Juan Vladimir de la; Castrillo, Antonio; Crawshaw, Dylan J.; Vidal Taboada, José Manuel; Saura Martí, Josep; Matalonga, Jonathan; Nunes Chini, Eduardo; Caelles Franch, Carme; Valledor Fernández, Annabel
Introduction: Macrophages abundantly express liver X receptors (LXRs), which are ligand-dependent transcription factors and sensors of several cholesterol metabolites. In response to agonists, LXRs promote the expression of key lipid homeostasis regulators. Cross talk between LXRs and inflammatory signals exists in a cell type- and gene-specific manner. A common feature in the macrophage response to inflammatory mediators is the induction of CCAAT/enhancer-binding protein beta (C/EBPβ), a master transcriptional regulator and lineage-determining transcription factor in monocytes/macrophages. Methods: Quantitative real-time PCR in control and C/EBPβ-deficient macrophages was used to explore the role of C/EBPβ in the cross talk between inflammatory mediators and the macrophage response to pharmacological LXR activation. The functional interaction between C/EBPβ and LXRs on selected genomic regions was further characterized by chromatin-immunoprecipitation (ChIP) and gene reporter studies. Results: Whereas inflammatory signaling repressed several LXR-regulated genes involved in lipid metabolism, these effects were conserved after deletion of C/EBPβ. In contrast, inflammatory mediators and LXRs synergistically induced the expression of the multifunctional protein CD38 in a C/EBPβ-dependent manner. C/EBPβ and LXRs bound to several regions with enhancer activity upstream and within the mouse Cd38 gene and their functional cooperation in macrophages required intact binding sites for LXR and C/EBPβ. Conclusion: This study reveals positive cross talk between C/EBPβ and LXRs during the macrophage inflammatory response, which selectively impacts CD38 expression.
A monoclonal antibody targeting a large surface of the receptor binding motif shows pan-neutralizing SARS-CoV-2 activity
(Nature Publishing Group, 2024-02-05) Campos Mata, Leire de; Trinité, Benjamin; Modrego Guillén, Andrea; Tejedor Vaquero, Sonia; Pradenas Saavedra, Edwards; Pons Grífols, Anna; Rodrigo Melero, Natalia; Carlero, Diego; Marfil, Silvia; Santiago, César; Raïch Regué, Dàlia; Bueno Carrasco, María Teresa; Tarrés Freixas, Ferran; Abancó, Ferran; Urrea, Victor; Izquierdo Useros, Nuria; Riveira Muñoz, Eva; Ballana, Ester; Pérez Maillo, Mónica; Vergara Alert, Júlia; Segalés, Joaquim; Carolis, Carlo; Arranz, Rocío; Blanco, Julià; Magri, Giuliana
Here we report the characterization of 17T2, a SARS-CoV-2 pan-neutralizing human monoclonal antibody isolated from a COVID-19 convalescent individual infected during the first pandemic wave. 17T2 is a class 1 VH1-58/κ3-20 antibody, derived from a receptor binding domain (RBD)-specific IgA+ memory B cell, with a broad neutralizing activity against former and new SARS-CoV-2 variants, including XBB.1.16 and BA.2.86 Omicron subvariants. Consistently, 17T2 demonstrates in vivo prophylactic and therapeutic activity against Omicron BA.1.1 infection in K18-hACE2 mice. Cryo-electron microscopy reconstruction shows that 17T2 binds the BA.1 spike with the RBD in "up" position and blocks the receptor binding motif, as other structurally similar antibodies do, including S2E12. Yet, unlike S2E12, 17T2 retains its neutralizing activity against all variants tested, probably due to a larger RBD contact area. These results highlight the impact of small structural antibody changes on neutralizing performance and identify 17T2 as a potential candidate for future clinical interventions.
A multi-site study on sex differences in cortical thickness in non-demented Parkinson’s disease
(Springer Nature, 2024-12-01) Oltra González, Javier; Segura i Fàbregas, Bàrbara; Strafella, Antonio P.; Eimeren, Thilo van; Díez Cirarda, María; Ibarretxe Bilbao, Naroa; Monté Rubio, Gemma C.; Ojeda, Natalia; Peña Lasa, Javier; Eggers, Carsten; Lucas Jiménez, Olaia; Ruppert, Marina C.; Sala Llonch, Roser; Theis, Hendrik; Uribe, Carme; Junqué i Plaja, Carme, 1955-; Factors sexuals en les malalties
Clinical, cognitive, and atrophy characteristics depending on sex have been previously reported in Parkinson’s disease (PD). However, though sex differences in cortical gray matter measures in early drugnaïvepatientshavebeendescribed,littleisknownaboutdifferencesincorticalthickness(CTh)as the disease advances. Our multi-site sample comprised 211 non-demented PD patients (64.45% males; mean age 65.58±8.44 years old; mean disease duration 6.42±5.11 years) and 86 healthy controls (50% males; mean age 65.49±9.33years old) with available T1-weighted 3T MRI datafrom four international research centers. Sex differences in regional mean CTh estimations were analyzed using generalized linear models. The relation of CTh in regions showing sex differences with age, disease duration, and age of onset was examined through multiple linear regression. PD males showedthinner cortex than PD females in six frontal (bilateral caudal middle frontal, bilateral superior frontal, left precentral and right pars orbitalis), three parietal (bilateral inferior parietal and left supramarginal), andonelimbicregion(right posterior cingulate).In PDmales,lowerCThvaluesinnine outoftenregionswereassociatedwithlongerdiseasedurationandolderage,whereasinPDfemales, lower CThwasassociatedwith older agebut with longer disease duration only in one region. Overall, male patients show a more widespread pattern of reduced CTh compared with female patients. Disease duration seems more relevant to explain reduced CTh in male patients, suggesting worse prognostic over time. Further studies should explore sex-specific cortical atrophy trajectories using large longitudinal multi-site data
RTP801 interacts with the tRNA ligase complex and dysregulates its RNA ligase activity in Alzheimer's disease
(Oxford University Press, 2024-10-14) Campoy Campos, Genís; Solana Balaguer, Júlia; Guisado Corcoll, Anna; Chicote González, Almudena; García Segura, Pol; Pérez Sisqués, Leticia; Gabriel Torres, Adrián; Canal de la Iglesia, Mercè; Molina Porcel, Laura; Fernández Irigoyen, Joaquín; Santamaría, Enrique; Ribas de Pouplana, Lluís; Alberch i Vié, Jordi, 1959-; Marti Puig, Eulalia; Giralt Torroella, Albert; Pérez Navarro, Esther; Malagelada Grau, Cristina
RTP801/REDD1 is a stress-responsive protein overexpressed in neurodegenerative diseases such as Alzheimer’s disease (AD) that contributes to cognitive deficits and neuroinflammation. Here, we found that RTP801 interacts with HSPC117, DDX1 and CGI-99, three members of the tRNA ligase complex (tRNA-LC), which ligates the excised exons of intron-containing tRNAs and the mRNA exons of the transcription factor XBP1 during the unfolded protein response (UPR). We also found that RTP801 modulates the mRNA ligase activity of the complex in vitro since RTP801 knockdown promoted XBP1 splicing and the expression of its transcriptional target, SEC24D. Conversely, RTP801 overexpression inhibited the splicing of XBP1. Similarly, in human AD postmortem hippocampal samples, where RTP801 is upregulated, we found that XBP1 splicing was dramatically decreased. In the 5xFAD mouse model of AD, silencing RTP801 expression in hippocampal neurons promoted Xbp1 splicing and prevented the accumulation of intron-containing pre-tRNAs. Finally, the tRNA-enriched fraction obtained from 5xFAD mice promoted abnormal dendritic arborization in cultured hippocampal neurons, and RTP801 silencing in the source neurons prevented this phenotype. Altogether, these results show that elevated RTP801 impairs RNA processing in vitro and in vivo in the context of AD and suggest that RTP801 inhibition could be a promising therapeutic approach.
Identification of Dhx15 as a Major Regulator of Liver Development, Regeneration, and Tumor Growth in Zebrafish and Mice
(MDPI, 2024-03-27) Portolés, Irene; Ribera, Jordi; Fernández Galán, Esther; Lecue Costas, Elena; Casals Mercadal, Gregori; Melgar Lesmes, Pedro; Fernández Varo, Guillermo; Boix i Ferrero, Loreto; Sanduzzi Zamparelli, Marco; Aishwarya, Veenu; Reig, María; Jiménez Povedano, Wladimiro; Morales Ruiz, Manuel
RNA helicase DHX15 plays a significant role in vasculature development and lung metastasis in vertebrates. In addition, several studies have demonstrated the overexpression of DHX15 in the context of hepatocellular carcinoma. Therefore, we hypothesized that this helicase may play a significant role in liver regeneration, physiology, and pathology. Dhx15 gene deficiency was generated by CRISPR/Cas9 in zebrafish and by TALEN-RNA in mice. AUM Antisense-Oligonucleotides were used to silence Dhx15 in wild-type mice. The hepatocellular carcinoma tumor induction model was generated by subcutaneous injection of Hepa 1-6 cells. Homozygous Dhx15 gene deficiency was lethal in zebrafish and mouse embryos. Dhx15 gene deficiency impaired liver organogenesis in zebrafish embryos and liver regeneration after partial hepatectomy in mice. Also, heterozygous mice presented decreased number and size of liver metastasis after Hepa 1-6 cells injection compared to wild-type mice. Dhx15 gene silencing with AUM Antisense-Oligonucleotides in wild-type mice resulted in 80% reduced expression in the liver and a significant reduction in other major organs. In addition, Dhx15 gene silencing significantly hindered primary tumor growth in the hepatocellular carcinoma experimental model. Regarding the potential use of DHX15 as a diagnostic marker for liver disease, patients with hepatocellular carcinoma showed increased levels of DHX15 in blood samples compared with subjects without hepatic affectation. In conclusion, Dhx15 is a key regulator of liver physiology and organogenesis, is increased in the blood of cirrhotic and hepatocellular carcinoma patients, and plays a key role in controlling hepatocellular carcinoma tumor growth and expansion in experimental models.
Preserved VPS13A distribution and expression in Huntington’s disease: Divergent mechanisms of action for similar movement disorders?
(Frontiers Media, 2024-06-05) García García, Esther; Carreras Caballé, Maria; Coll Manzano, Albert; Ramón Lainez, Alba; Besa Selva, Gisela; Pérez Navarro, Esther; Malagelada Grau, Cristina; Alberch i Vié, Jordi; Masana Nadal, Mercè; Rodríguez Allué, Manuel José
VPS13A disease and Huntington’s disease (HD) are two basal ganglia disorders that may be difficult to distinguish clinically because they have similar symptoms, neuropathological features, and cellular dysfunctions with selective degeneration of the medium spiny neurons of the striatum. However, their etiology is different. VPS13A disease is caused by a mutation in the VPS13A gene leading to a lack of protein in the cells, while HD is due to an expansion of CAG repeat in the huntingtin (Htt) gene, leading to aberrant accumulation of mutant Htt. Considering the similarities of both diseases regarding the selective degeneration of striatal medium spiny neurons, the involvement of VPS13A in the molecular mechanisms of HD pathophysiology cannot be discarded. We analyzed the VPS13A distribution in the striatum, cortex, hippocampus, and cerebellum of a transgenic mouse model of HD. We also quantified the VPS13A levels in the human cortex and putamen nucleus; and compared data on mutant Htt-induced changes in VPS13A expression from differential expression datasets. We found that VPS13A brain distribution or expression was unaltered in most situations with a decrease in the putamen of HD patients and small mRNA changes in the striatum and cerebellum of HD mice. We concluded that the selective susceptibility of the striatum in VPS13A disease and HD may be a consequence of disturbances in different cellular processes with convergent molecular mechanisms already to be elucidated.
Palmitoylcarnitine impairs immunity in decompensated cirrhosis
(Elsevier, 2024-08-22) Wei Zhang, Ingrid; Sánchez Rodríguez, María Belén; López Vicario, Cristina; Casulleras, Mireia; Duran Güell, Marta; Flores Costa, Roger; Aguilar, Ferran; Rothe, Michael; Segalés, Paula; García Ruiz, Carmen; Fernández Checa Torres, José Carlos; Trebicka, Jonel; Arroyo, Vicente; Clària i Enrich, Joan
Background & aims: In patients with cirrhosis, acute decompensation (AD) correlates with a hyperinflammatory state driven by mitochondrial dysfunction, which is a significant factor in the progression toward acute-on-chronic liver failure (ACLF). Elevated circulating levels of acylcarnitine, indicative of mitochondrial dysfunction, are predictors of mortality in ACLF patients. Our hypothesis posits that acylcarnitines not only act as biomarkers, but also actively exert detrimental effects on circulating immune cells. Methods: Plasma acylcarnitine levels were measured in 20 patients with AD cirrhosis and 10 healthy individuals. The effects of selected medium- and long-chain acylcarnitines on mitochondrial function were investigated in peripheral leucocytes from healthy donors by determining mitochondrial membrane potential (Δψm) and mitochondrial respiration using the JC-1 dye and Agilent Seahorse XF technology. Changes regarding mitochondrial ultrastructure and redox systems were assessed by transmission electron microscopy and gene and protein expression analysis. Results: Plasma levels of several acylcarnitine species were significantly elevated in patients with AD cirrhosis compared with healthy individuals, alongside increased levels of inflammatory mediators (cytokines and chemokines). Notably, the long-chain acylcarnitine palmitoylcarnitine (C16:0-carnitine, 1.51-fold higher, p = 0.0059) impaired Δψm and reduced the spare respiratory capacity of peripheral mononuclear leucocytes. Additionally, C16:0-carnitine induced mitochondrial oxidative stress, suppressed the expression of the antioxidant gene HMOX1, and increased CXCL8 expression and IL-8 release. Etomoxir, which blocks acylcarnitine entry into the mitochondria, reversed the suppression of HMOX1. Similarly, trimetazidine, a fatty acid beta-oxidation inhibitor, prevented C16:0-carnitine-induced CXCL8 expression. Importantly, oxidative stress and Δψm impairment caused by C16:0-carnitine were less severe in the presence of albumin, a standard therapy for AD cirrhosis. Conclusions: Our findings suggest that long-chain acylcarnitines induce mitochondrial injury in immune cells, thereby contributing to the development of immune dysfunction associated with cirrhosis. Impact and implications: Patients with acute decompensation of cirrhosis and acute-on-chronic liver failure (ACLF) display a systemic hyperinflammatory state and leukocyte mitochondrial dysfunction. We discovered that apart from being increased in the circulation of these patients, the long-chain palmitoylcarnitine is able to elicit cytokine secretion paired with mitochondrial dysfunction in leukocytes from healthy donors. In particular, we show that inhibiting the metabolism of palmitoylcarnitine could reverse these detrimental effects. Our findings underline the importance of immunometabolism as a treatment target in patients with acute decompensation of cirrhosis and ACLF.
Reciprocal Negative Cross-Talk between Liver X Receptors (LXRs) and STAT1: Effects on IFN-gamma-Induced Inflammatory Responses and LXR-Dependent Gene Expression
(American Association of Immunologists, 2013-06-15) Pascual Garcia, Monica; Rué Cabré, Laura; León Moreno, Theresa Elizabeth; Julve, Josep; Carbó, José M.; Matalonga, Jonathan; Auer, Herbert; Celada Cotarelo, Antonio; Escola-Gil, Joan Carles; Steffensen, Knut R.; Pérez Navarro, Esther; Valledor Fernandez, Annabel
Liver X receptors (LXRs) exert key functions in lipid homeostasis and in control of inflammation. In this study we have explored the impact of LXR activation on the macrophage response to the endogenous inflammatory cytokine IFN-γ. Transcriptional profiling studies demonstrate that ∼38% of the IFN-γ–induced transcriptional response is repressed by LXR activation in macrophages. LXRs also mediated inhibitory effects on selected IFN-γ–induced genes in primary microglia and in a model of IFN-γ–induced neuroinflammation in vivo. LXR activation resulted in reduced STAT1 recruitment to the promoters tested in this study without affecting STAT1 phosphorylation. A closer look into the mechanism revealed that SUMOylation of LXRs, but not the presence of nuclear receptor corepressor 1, was required for repression of the NO synthase 2 promoter. We have also analyzed whether IFN-γ signaling exerts reciprocal effects on LXR targets. Treatment with IFN-γ inhibited, in a STAT1-dependent manner, the LXR-dependent upregulation of selective targets, including ATP-binding cassette A1 (ABCA1) and sterol response element binding protein 1c. Downregulation of ABCA1 expression correlated with decreased cholesterol efflux to apolipoprotein A1 in macrophages stimulated with IFN-γ. The inhibitory effects of IFN-γ on LXR signaling did not involve reduced binding of LXR/retinoid X receptor heterodimers to target gene promoters. However, overexpression of the coactivator CREB-binding protein/p300 reduced the inhibitory actions of IFN-γ on the Abca1 promoter, suggesting that competition for CREB-binding protein may contribute to STAT1-dependent downregulation of LXR targets. The results from this study suggest an important level of bidirectional negative cross-talk between IFN-γ/STAT1 and LXRs with implications both in the control of IFN-γ–mediated immune responses and in the regulation of lipid metabolism.
Liver X Receptors Inhibit Macrophage Proliferation through Downregulation of Cyclins D1 and B1 and Cyclin-Dependent Kinases 2 and 4
(American Association of Immunologists, 2011-04-15) Pascual García, Mónica; Carbó, José M.; León Moreno, Theresa Elizabeth; Matalonga, Jonathan; Out, Ruud; Van Berkel, Theo; Sarrias, Maria Rosa; Lozano, Francisco; Celada Cotarelo, Antonio; Valledor Fernández, Annabel
Macrophages serve essential functions as regulators of immunity and homeostasis, and their proliferation contributes to pathogenesis of certain disorders. In this report, we show that induction of macrophage proliferation by the growth factor M-CSF is negatively modulated by agonists that activate the nuclear receptor liver X receptor (LXR), both in vitro and in vivo. Both isoforms LXR α and β are involved in the antiproliferative actions of LXR ligands in macrophages. In contrast, M-CSF does not exert negative effects on LXR-mediated gene expression. Treatment with LXR agonists results in the accumulation of macrophages in the G₀/G₁ phase of the cell cycle without affecting ERK-1/2 phosphorylation. The use of small interfering RNA or genetically modified mice revealed that, in contrast to other cellular models, functional expression of either the cyclin-dependent kinase inhibitor p27KIP1 or the cholesterol transporters ATP-binding cassette A1 or ATP-binding cassette G1 was not required for the antiproliferative effects of LXR agonists in macrophages. Western blot analysis revealed that protein expression of key molecules that regulate progression through the cell cycle, such as cyclins D1 and B1 and cyclin-dependent kinases 2 and 4, was downregulated upon LXR activation. These observations suggest a role for LXR agonists in limiting macrophage proliferative responses associated to pathogenic disorders.
Hippocampal Pyk2 regulates specific social skills: Implications for schizophrenia
(Elsevier, 2024-03-27) López Molina, Laura; Sancho Balsells, Anna; Al-Massadi, Omar; Montalban, Enrica; Alberch i Vié, Jordi, 1959-; Arranz, Belém; Girault, Jean-Antoine; Giralt Torroella, Albert
Pyk2 has been shown previously to be involved in several psychological and cognitive alterations related to stress, Huntington's disease, and Alzheimer's disease. All these disorders are accompanied by different types of impairments in sociability, which has recently been linked to improper mitochondrial function. We hypothesize that Pyk2, which regulates mitochondria, could be associated with the regulation of mitochondrial dynamics and social skills. In the present manuscript, we report that a reduction of Pyk2 levels in mouse pyramidal neurons of the hippocampus decreased social dominance and aggressivity. Furthermore, social interactions induced robust Pyk2-dependent hippocampal changes in several oxidative phosphorylation complexes. We also observed that Pyk2 levels were increased in the CA1 pyramidal neurons of schizophrenic subjects, occurring alongside changes in different direct and indirect regulators of mitochondrial function including DISC1 and Grp75. Accordingly, overexpressing Pyk2 in hippocampal CA1 pyramidal cells mimicked some specific schizophrenia-like social behaviors in mice. In summary, our results indicate that Pyk2 might play a role in regulating specific social skills likely via mitochondrial dynamics and that there might be a link between Pyk2 levels in hippocampal neurons and social disturbances in schizophrenia.
Liver-targeted nanoparticles delivering nitric oxide reduce portal hypertension in cirrhotic rats
(Elsevier Masson SAS, 2024-01-13) Perramón, Meritxell; Navalón López, María; Fernández Varo, Guillermo; Moreno Lanceta, Alazne; García Pérez, Rocío; Faneca, Joana; López Moya, Mario; Fornaguera, Cristina; García Villoria, Judit; Morales Ruiz, Manuel; Melgar-Lesmes, Pedro; Borrós, Salvador; Jiménez Povedano, Wladimiro
Nitric oxide (NO) is a small vasodilator playing a key role in the pathogenesis of portal hypertension. Here, we assessed the potential therapeutic effect of a NO donor targeted to the liver by poly(beta-amino ester) nanoparticles (pBAE NPs) in experimental cirrhosis. Retinol-functionalized NO donor pBAE NPs (Ret pBAE NPs) were synthetized with the aim of actively targeting the liver. Administration of Ret pBAE NPs resulted in uptake and transfection by the liver and spleen. NPs were not found in other organs or the systemic circulation. Treatment with NO donor Ret pBAE NPs (30 mg/ kg body weight) significantly decreased aspartate aminotransferase, lactate dehydrogenase and portal pressure (9.75 ± 0.64 mmHg) compared to control NPs (13.4 ± 0.53 mmHg) in cirrhotic rats. There were no effects on mean arterial pressure and cardiac output. Liver-targeted NO donor NPs reduced collagen fibers and steatosis, activation of hepatic stellate cells and mRNA expression of profibrogenic and proinflammatory genes. Finally, Ret pBAE NPs displayed efficient transfection in human liver slices. Overall, liver-specific NO donor NPs effectively target the liver and mitigated inflammation and portal hypertension in cirrhotic rats. The use of Ret pBAE may prove to be an effective therapeutic strategy to treat advanced liver disease.
Nutritional profile of the diet according to circadian clock genes in the European Prospective Investigation into Cancer and Nutrition (EPIC) chronodiet study
(Elsevier BV, 2025-04-25) Molina Montes, Esther; Rodríguez Barranco, Miguel; Alcalá Santiago, Ángela; Gálvez Navas, José María; Huerta Castaño, José María; Amiano, Pilar; Lasheras, Cristina; Moreno Iribas, Conchi; Jiménez Zabala, Ana; Chirlaque, María Dolores; Gasque, Alba; Luján Barroso, Leila; Agudo, Antonio; Jakszyn, Paula; Ramón Quirós, José; Sánchez Pérez, María José
Background & aims: Circadian rhythms seem to impact both dietary intake and metabolism, depending on the individual's chronotype. We aimed to explore whether the nutritional composition of meals throughout the day is influenced by genetics linked to the circadian clock and chronotype within the European Prospective Investigation into Cancer and Nutrition (EPIC) chronodiet study. Methods: The study population comprised 3,183 subjects with information on diet and twelve genetic variants of six genes (PER1, PER2, PER3, CRY1, NR1D1, CLOCK). The associations between the variants with chrononutrition variables (macronutrients and serving sizes of each meal) were evaluated using linear regression, considering an additive genetic model, and adjusting for sex, age and center, among others. The beta coefficients, 95 % confidence intervals (CI), and p-values corrected for multiple comparisons were estimated. A genetic risk score (GRS) that was associated to the evening/late chronotype as well as overweight/obesity in a previous study, the chronotype-GRS, was tested for its association with chrononutrition variables. Results: The nutritional profile of the diet differed according to the individual's chronotype, with evening/late chronotypes exhibiting an unbalanced intake during breakfast and dinner compared to the intermediate and early chronotypes (e.g., percentage of fats consumed at breakfast relative to the total fat intake: 13 % and 9 %, respectively). However, significant differences were not encountered by the chronotype-GRS. In multivariate analyses, individual associations between the genetic variants and the nutrients revealed some nominal associations (e.g., rs1801260 and rs2070062 with carbohydrates at breakfast: beta = -0.06 to 0.08). Higher scorings of the chronotype-GRS were inversely associated with the intake of proteins and carbohydrates (beta = -0.46 and -0.41; nominal p-value<0.006; corrected = 0.25) during breakfast. Also, there was an inverse association between the chronotype-GRS and the breakfast's portion size (beta = -0.3; nominal p-value = 0.03; corrected = 0.1). Conclusions: Genetic susceptibility to an evening-like chronotype prone to overweight/obesity seems to be associated with a smaller serving size during breakfast, with lower protein and carbohydrate content. (c) 2025 The Author(s). Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
Motor skill learning modulates striatal extracellular vesicles’ content in a mouse model of Huntington’s disease
(BioMed Central, 2024-06-11) Solana Balaguer, Júlia; Garcia-Segura, Pol; Campoy Campos, Genís; Chicote González, Almudena; Fernández Irigoyen, Joaquín; Santamaría Hernández, Esther; Pérez Navarro, Esther; Masana Nadal, Mercè; Alberch i Vié, Jordi; Malagelada Grau, Cristina
Huntington’s disease (HD) is a neurological disorder caused by a CAG expansion in the Huntingtin gene (HTT). HD pathology mostly affects striatal medium-sized spiny neurons and results in an altered cortico-striatal function. Recent studies report that motor skill learning, and cortico-striatal stimulation attenuate the neuropathology in HD, resulting in an amelioration of some motor and cognitive functions. During physical training, extracellular vesicles (EVs) are released in many tissues, including the brain, as a potential means for inter-tissue communication. To investigate how motor skill learning, involving acute physical training, modulates EVs crosstalk between cells in the striatum, we trained wild-type (WT) and R6/1 mice, the latter with motor and cognitive deficits, on the accelerating rotarod test, and we isolated their striatal EVs. EVs from R6/1 mice presented alterations in the small exosome population when compared to WT. Proteomic analyses revealed that striatal R6/1 EVs recapitulated signaling and energy deficiencies present in HD. Motor skill learning in R6/1 mice restored the amount of EVs and their protein content in comparison to naïve R6/1 mice. Furthermore, motor skill learning modulated crucial pathways in metabolism and neurodegeneration. All these data provide new insights into the pathogenesis of HD and put striatal EVs in the spotlight to understand the signaling and metabolic alterations in neurodegenerative diseases. Moreover, our results suggest that motor learning is a crucial modulator of cell-to-cell communication in the striatum.
Discovery of the first PD-1 ligand encoded by a pathogen
(Frontiers Media, 2022-09-13) Martínez Vicente, Pablo; Poblador Bonet, Francesc; Leitner, Judith; Farré Marimon, Domènec; Steinberger, Peter; Engel Rocamora, Pablo; Angulo Aguado, Ana
Large double-stranded DNA viruses deploy multiple strategies to subvert host immune defenses. Some of these tactics are mediated by viral gene products acquired by horizontal gene transfer from the corresponding hosts and shaped throughout evolution. The programmed death-1 (PD-1) receptor and its ligands, PD-L1 and PD-L2, play a pivotal role attenuating T-cell responses and regulating immune tolerance. In this study, we report the first functional PD-L1 homolog gene (De2) found in a pathogen. De2, captured by a gherpesvirus from its host during co-evolution around 50 million years ago, encodes a cell-surface glycoprotein that interacts with high affinity and stability with host PD-1. We also find that mutations evolved by the viral protein result in a significant loss of its ability to interact in cis with CD80, an interaction that for PD-L1:CD80 has been reported to block PD-1 inhibitory pathways. Furthermore, we demonstrate that the viral protein strongly inhibits T-cell signaling. Our observations suggest that PD-L1 homologs may enable viruses to evade T cell responses, favor their replication, and prevent excessive tissue damage. Altogether, our findings reveal a novel viral immunosuppressive strategy and highlight the importance of the modulation of the PD-1/PD-L1 axis during viral infections.
Standardization of Workflow and Flow Cytometry Panels for Quantitative Expression Profiling of Surface Antigens on Blood Leukocyte Subsets: An HCDM CDMaps Initiative
(Frontiers Media, 2022-02-11) Kuzilková, Daniela; Puñet Ortiz, Joan; Aui, Pei M.; Fernández, Javier; Fiser, Karel; Engel Rocamora, Pablo; van Zelm, Menno C.; Kalina, Tomas
Background: The Human Cell Differentiation Molecules (HCDM) organizes Human Leukocyte Differentiation Antigen (HLDA) workshops to test and name clusters of antibodies that react with a specific antigen. These cluster of differentiation (CD) markers have provided the scientific community with validated antibody clones, consistent naming of targets and reproducible identification of leukocyte subsets. Still, quantitative CD marker expression profiles and benchmarking of reagents at the single-cell level are currently lacking. Objective: To develop a flow cytometric procedure for quantitative expression profiling of surface antigens on blood leukocyte subsets that is standardized across multiple research laboratories. Methods: A high content framework to evaluate the titration and reactivity of Phycoerythrin (PE)-conjugated monoclonal antibodies (mAbs) was created. Two flow cytometry panels were designed: an innate cell tube for granulocytes, dendritic cells, monocytes, NK cells and innate lymphoid cells (12-color) and an adaptive lymphocyte tube for naive and memory B and T cells, including TCRγδ+, regulatory-T and follicular helper T cells (11-color). The potential of these 2 panels was demonstrated via expression profiling of selected CD markers detected by PE-conjugated antibodies and evaluated using 561 nm excitation. Results: Using automated data annotation and dried backbone reagents, we reached a robust workflow amenable to processing hundreds of measurements in each experiment in a 96-well plate format. The immunophenotyping panels enabled discrimination of 27 leukocyte subsets and quantitative detection of the expression of PE-conjugated CD markers of interest that could quantify protein expression above 400 units of antibody binding capacity. Expression profiling of 4 selected CD markers (CD11b, CD31, CD38, CD40) showed high reproducibility across centers, as well as the capacity to benchmark unique clones directed toward the same CD3 antigen. Conclusion: We optimized a procedure for quantitative expression profiling of surface antigens on blood leukocyte subsets. The workflow, bioinformatics pipeline and optimized flow panels enable the following: 1) mapping the expression patterns of HLDA-approved mAb clones to CD markers; 2) benchmarking new antibody clones to established CD markers; 3) defining new clusters of differentiation in future HLDA workshops.
SIRPα - CD47 axis regulates dendritic cell-T cell interactions and TCR activation during T cell priming in spleen.
(Public Library of Science (PLoS), 2022-04-12) Autio, Anu; Wang, Huan; Velázquez, Francisco; Newton, Gail; Parkos, Charles A.; Engel Rocamora, Pablo; Engelbertsen, Daniel; Lichtman, Andrew H.; Luscinskas, Francis W.
The SIRPα-CD47 axis plays an important role in T cell recruitment to sites of immune reaction and inflammation but its role in T cell antigen priming is incompletely understood. Employing OTII TCR transgenic mice bred to Cd47-/- (Cd47KO) or SKI mice, a knock-in transgenic animal expressing non-signaling cytoplasmic-truncated SIRPα, we investigated how the SIRPα-CD47 axis contributes to antigen priming. Here we show that adoptive transfer of Cd47KO or SKI Ova-specific CD4+ T cells (OTII) into Cd47KO and SKI recipients, followed by Ova immunization, elicited reduced T cell division and proliferation indices, increased apoptosis, and reduced expansion compared to transfer into WT mice. We confirmed prior reports that splenic T cell zone, CD4+ conventional dendritic cells (cDCs) and CD4+ T cell numbers were reduced in Cd47KO and SKI mice. We report that in vitro derived DCs from Cd47KO and SKI mice exhibited impaired migration in vivo and exhibited reduced CD11c+ DC proximity to OTII T cells in T cell zones after Ag immunization, which correlates with reduced TCR activation in transferred OTII T cells. These findings suggest that reduced numbers of CD4+ cDCs and their impaired migration contributes to reduced T cell-DC proximity in splenic T cell zone and reduced T cell TCR activation, cell division and proliferation, and indirectly increased T cell apoptosis.
Deconstructing Complexity: A Computational Topology Approach to Trajectory Inference in the Human Thymus with TVIBLINDI
(eLife Sciences, 2025-04-23) Stuchly, Jan; Novak, David; Brdickova, Nadezda; Hadlova, Petra; Iksi, Ahmad; Kuzilkova, Daniela; Svaton, Michael; Saad, George- Alehandro; Engel Rocamora, Pablo; Luche, Herve; Sousa, Ana E.; Almeida, Afonso R. M.; Kalina, Tomas
Understanding complex, organ-level single-cell datasets represents a formidable interdisciplinary challenge. This study aims to describe developmental trajectories of thymocytes and mature T cells. We developed tviblindi, a trajectory inference algorithm that integrates several autonomous modules - pseudotime inference, random walk simulations, real-time topological classification using persistent homology, and autoencoder-based 2D visualization using the vaevictis algorithm. This integration facilitates interactive exploration of developmental trajectories, revealing not only the canonical CD4 and CD8 development but also offering insights into checkpoints such as TCRβ selection and positive/negative selection. Furthermore, tviblindi allowed us to thoroughly characterize thymic regulatory T cells,tracing their development passed the negative selection stage to mature thymic regulatory T cells. At the very end of the developmental trajectory we discovered a previously undescribed subpopulation of thymic regulatory T cells. Experimentally, we confirmed its extensive proliferation history and an immunophenotype characteristic of activated and recirculating cells. tviblindi represents a new class of methods that is complementary to fully automated trajectory inference tools. It offers a semi-automated tool that leverages features derived from data in an unbiased and mathematically rigorous manner. These features include pseudotime, homology classes, and appropriate low-dimensional representations. These features can be integrated with expert knowledge to formulate hypotheses regarding the underlying dynamics, tailored to the specific trajectory or biological process under investigation.
Novel selective strategies targeting the BCL-2 family to enhance clinical efficacy in ALK-rearranged non-small cell lung cancer
(Springer Science and Business Media LLC, 2025-03-20) Martín, Fernando; Alcon, Clara; Marín, Elba; Morales Sánchez, Paula; Manzano Muñoz, Albert; Diaz, Sherley; García López, Mireia; Samitier i Martí, Josep; Lu, Albert; Villanueva Garatachea, Alberto; Reguart, Noemí; Teixido Febrero, Cristina; Montero Boronat, Joan
ALK (anaplastic lymphoma kinase) rearrangements represent the third most predominant driver oncogene in non-small cell lung cancer (NSCLC). Although ALK inhibitors are the tyrosine kinase inhibitors (TKIs) with the longest survival rates in lung cancer, the complex systemic clinical evaluation and the apoptotic cell death evasion of drug-tolerant persister (DTP) cancer cells may limit their therapeutic response. We found that dynamic BH3 profiling (DBP) presents an excellent predictive capacity to ALK-TKIs, that would facilitate their use in a clinical setting and complementing the readout of standard diagnostic assays. In addition, we revealed novel acute adaptive mechanisms in response to ALK inhibitors in cell lines and patient-derived tumor cells. Consistently, all our cell models confirmed a rapid downregulation of the sensitizer protein NOXA, leading to dependence on the anti-apoptotic protein MCL-1 after treatment with ALK-TKIs. In some cases, the anti-apoptotic protein BCL-xL may contribute equally to this anti-apoptotic response. Importantly, these acute dependencies could be prevented with BH3 mimetics in vitro and in vivo, blocking tumor adaptation to treatment. Finally, we also demonstrated how dual reactivation of PI3K/AKT and MAPK signaling pathways can impair lorlatinib response, which could be overcome with specific inhibitors of both signaling pathways. In conclusion, our findings propose several therapeutic combinations that should be explored in future clinical trials to enhance ALK inhibitors efficacy and improve the clinical response in a broad NSCLC patient population.
Validation of an LC–HRMS Method for Quantifying Indoxyl Sulfate and p-Cresyl Sulfate in Human Serum
(MDPI, 2025-02-08) Rodríguez García, María; Martínez Martín, Irene; Aliart, Irene; Sainz de Medrano, Jaime I.; Rico, Nayra; Escudero Saiz, Víctor Joaquín; Maduell, Francisco; Morales Ruiz, Manuel; Casals Mercadal, Gregori
Accurate quantification of indoxyl sulfate (IndS) and p-cresyl sulfate (pCS) is essential for understanding their role in chronic kidney disease (CKD) progression and for developing strategies to mitigate their harmful effects, including cardiovascular morbidity and renal fibrosis. Advances in liquid chromatography-high-resolution mass spectrometry (LC-HRMS) enable the integration of powerful diagnostic tools into clinical laboratories. Along with accurate quantification, precise mass measurements allow for untargeted compound identification. Methods: An LC-HRMS was validated for quantifying IndS and pCS in human serum, following EMA guidelines. The method involved protein precipitation with methanol, micro-LC for chromatographic separation, and detection based on accurate mass, with simultaneous high-resolution full-scan acquisition. Clinical samples from patients with varying degrees of renal insufficiency and samples obtained before and after hemodiafiltration were analyzed. Results: The method demonstrated acceptable linearity, precision, and accuracy. The measurement range for both analytes was from 100 to 40,000 ng/mL. Serum levels of IndS and pCS correlated with decreased renal function. After hemodiafiltration, there was a significant reduction of IndS (50%) and pCS (43%). Simultaneous untargeted analysis allowed to identify metabolites significantly underexpressed after hemodiafiltration. Conclusions: An accurate LC-HRMS method was validated for the quantification of IndS and pCS serum levels in patients with CKD, providing insights into toxin dynamics and enabling untargeted metabolic evaluation.

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