RESUMEN
Lipid and cholesterol metabolism play a crucial role in tumor cell behavior and in shaping the tumor microenvironment. In particular, enzymatic and non-enzymatic cholesterol metabolism, and derived metabolites control dendritic cell (DC) functions, ultimately impacting tumor antigen presentation within and outside the tumor mass, dampening tumor immunity and immunotherapeutic attempts. The mechanisms accounting for such events remain largely to be defined. Here we perturbed (oxy)sterol metabolism genetically and pharmacologically and analyzed the tumor lipidome landscape in relation to the tumor-infiltrating immune cells. We report that perturbing the lipidome of tumor microenvironment by the expression of sulfotransferase 2B1b crucial in cholesterol and oxysterol sulfate synthesis, favored intratumoral representation of monocyte-derived antigen-presenting cells, including monocyte-DCs. We also found that treating mice with a newly developed antagonist of the oxysterol receptors Liver X Receptors (LXRs), promoted intratumoral monocyte-DC differentiation, delayed tumor growth and synergized with anti-PD-1 immunotherapy and adoptive T cell therapy. Of note, looking at LXR/cholesterol gene signature in melanoma patients treated with anti-PD-1-based immunotherapy predicted diverse clinical outcomes. Indeed, patients whose tumors were poorly infiltrated by monocytes/macrophages expressing LXR target genes showed improved survival over the course of therapy. Thus, our data support a role for (oxy)sterol metabolism in shaping monocyte-to-DC differentiation, and in tumor antigen presentation critical for responsiveness to immunotherapy. The identification of a new LXR antagonist opens new treatment avenues for cancer patients.
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Melanoma , Monocitos , Ratones , Animales , Monocitos/metabolismo , Diferenciación Celular , Colesterol/metabolismo , Presentación de Antígeno , Células Dendríticas/metabolismo , Microambiente TumoralRESUMEN
Hepatic metastases are a poor prognostic factor of colorectal carcinoma (CRC) and new strategies to reduce the risk of liver CRC colonization are highly needed. Herein, we used mouse models of hepatic metastatization to demonstrate that the continuous infusion of therapeutic doses of interferon-alpha (IFNα) controls CRC invasion by acting on hepatic endothelial cells (HECs). Mechanistically, IFNα promoted the development of a vascular antimetastatic niche characterized by liver sinusoidal endothelial cells (LSECs) defenestration extracellular matrix and glycocalyx deposition, thus strengthening the liver vascular barrier impairing CRC trans-sinusoidal migration, without requiring a direct action on tumor cells, hepatic stellate cells, hepatocytes, or liver dendritic cells (DCs), Kupffer cells (KCs) and liver capsular macrophages (LCMs). Moreover, IFNα endowed LSECs with efficient cross-priming potential that, along with the early intravascular tumor burden reduction, supported the generation of antitumor CD8+ T cells and ultimately led to the establishment of a protective long-term memory T cell response. These findings provide a rationale for the use of continuous IFNα therapy in perioperative settings to reduce CRC metastatic spreading to the liver.
Colorectal cancer remains one of the most widespread and deadly cancers worldwide. Poor health outcomes are usually linked to diseased cells spreading from the intestine to create new tumors in the liver or other parts of the body. Treatment involves surgically removing the initial tumors in the bowel, but patient survival could be improved if, in parallel, their immune system was 'boosted' to destroy cancer cells before they can form other tumors. Interferon alpha is a small protein which helps to coordinate how the immune system recognizes and deactivates foreign agents and cancerous cells. It has recently been trialed as a colorectal cancer treatment to prevent tumors from spreading to the liver, but only with limited success. This partly because interferon-alpha is usually administered in high and pulsed doses, which cause severe side effects through the body. Instead, Tran, Ferreira, Alvarez-Moya et al. aimed to investigate whether continuously delivering lower amounts of the drug could be a better approach. This strategy was tested on mice in which colorectal cancer cells had been implanted into the wall of the large intestine. Continuous administration minimized the risk of the implanted cancer cells spreading to the liver while also creating fewer side effects. The team was able to identify an optimum delivery strategy by varying how much interferon-alpha the animals received and when. Further experiments also revealed a new mechanism by which interferon-alpha prevented the spread of colorectal cancer. Upon receiving continuous doses of the drug, a group of liver cells started to generate a physical barrier which stopped cancer cells from being able to invade the organ. The treatment also promoted long-term immune responses that targeted diseased cells while being safe for healthy tissues. If confirmed in clinical trials, these results suggest that colorectal patients undergoing tumor removal surgery may benefit from also receiving interferon-alpha through continuous delivery.
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Neoplasias Colorrectales , Interferón-alfa , Animales , Ratones , Células Endoteliales/patología , Linfocitos T CD8-positivos , Hígado , Hepatocitos , Neoplasias Colorrectales/patologíaRESUMEN
Kupffer cells (KCs) are highly abundant, intravascular, liver-resident macrophages known for their scavenger and phagocytic functions. KCs can also present antigens to CD8+ T cells and promote either tolerance or effector differentiation, but the mechanisms underlying these discrepant outcomes are poorly understood. Here, we used a mouse model of hepatitis B virus (HBV) infection, in which HBV-specific naive CD8+ T cells recognizing hepatocellular antigens are driven into a state of immune dysfunction, to identify a subset of KCs (referred to as KC2) that cross-presents hepatocellular antigens upon interleukin-2 (IL-2) administration, thus improving the antiviral function of T cells. Removing MHC-I from all KCs, including KC2, or selectively depleting KC2 impaired the capacity of IL-2 to revert the T cell dysfunction induced by intrahepatic priming. In summary, by sensing IL-2 and cross-presenting hepatocellular antigens, KC2 overcome the tolerogenic potential of the hepatic microenvironment, suggesting new strategies for boosting hepatic T cell immunity.
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Presentación de Antígeno/inmunología , Linfocitos T CD8-positivos/inmunología , Reactividad Cruzada/inmunología , Interleucina-2/inmunología , Macrófagos del Hígado/inmunología , Animales , Hepatitis B/inmunología , Tolerancia Inmunológica/inmunología , Ratones , Ratones TransgénicosRESUMEN
The responses of CD8+ T cells to hepatotropic viruses such as hepatitis B range from dysfunction to differentiation into effector cells, but the mechanisms that underlie these distinct outcomes remain poorly understood. Here we show that priming by Kupffer cells, which are not natural targets of hepatitis B, leads to differentiation of CD8+ T cells into effector cells that form dense, extravascular clusters of immotile cells scattered throughout the liver. By contrast, priming by hepatocytes, which are natural targets of hepatitis B, leads to local activation and proliferation of CD8+ T cells but not to differentiation into effector cells; these cells form loose, intravascular clusters of motile cells that coalesce around portal tracts. Transcriptomic and chromatin accessibility analyses reveal unique features of these dysfunctional CD8+ T cells, with limited overlap with those of exhausted or tolerant T cells; accordingly, CD8+ T cells primed by hepatocytes cannot be rescued by treatment with anti-PD-L1, but instead respond to IL-2. These findings suggest immunotherapeutic strategies against chronic hepatitis B infection.
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Linfocitos T CD8-positivos/inmunología , Reactividad Cruzada/inmunología , Virus de la Hepatitis B/inmunología , Hepatocitos/inmunología , Hepatocitos/virología , Animales , Antígeno B7-H1/antagonistas & inhibidores , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/metabolismo , Diferenciación Celular , Cromatina/metabolismo , Femenino , Hepatitis B/tratamiento farmacológico , Hepatitis B/inmunología , Hepatitis B/virología , Humanos , Tolerancia Inmunológica , Interleucina-2/inmunología , Interleucina-2/uso terapéutico , Macrófagos del Hígado/inmunología , Activación de Linfocitos , Masculino , Ratones , Transcriptoma/genéticaRESUMEN
Altered macrophage infiltration upon tissue damage results in inadequate healing due to inappropriate remodeling and stem cell recruitment and differentiation. We investigated in vivo whether cells of endothelial origin phenotypically change upon heterotopic ossification induction and whether infiltration of innate immunity cells influences their commitment and alters the ectopic bone formation. Liposome-encapsulated clodronate was used to assess macrophage impact on endothelial cells in the skeletal muscle upon acute damage in the ECs specific lineage-tracing Cdh5CreERT2:R26REYFP/dtTomato transgenic mice. Macrophage depletion in the injured skeletal muscle partially shifts the fate of ECs toward endochondral differentiation. Upon ectopic stimulation of BMP signaling, monocyte depletion leads to an enhanced contribution of ECs chondrogenesis and to ectopic bone formation, with increased bone volume and density, that is reversed by ACVR1/SMAD pathway inhibitor dipyridamole. This suggests that macrophages contribute to preserve endothelial fate and to limit the bone lesion in a BMP/injury-induced mouse model of heterotopic ossification. Therefore, alterations of the macrophage-endothelial axis may represent a novel target for molecular intervention in heterotopic ossification.
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Condrogénesis , Células Endoteliales/fisiología , Macrófagos/inmunología , Monocitos/inmunología , Músculo Esquelético/patología , Osificación Heterotópica/inmunología , Animales , Ratones Transgénicos , Músculo Esquelético/fisiologíaRESUMEN
Achaete-scute homolog 1 gene (ASCL1) is a gene classifier for the proneural (PN) transcriptional subgroup of glioblastoma (GBM) that has a relevant role in the neuronal-like differentiation of GBM cancer stem cells (CSCs) through the activation of a PN gene signature. Besides prototypical ASCL1 PN target genes, the molecular effectors mediating ASCL1 function in regulating GBM differentiation and, most relevantly, subgroup specification are currently unknown. Here we report that ASCL1 not only promotes the acquisition of a PN phenotype in CSCs by inducing a glial-to-neuronal lineage switch but also concomitantly represses mesenchymal (MES) features by directly downregulating the expression of N-Myc downstream-regulated gene 1 (NDRG1), which we propose as a novel gene classifier of MES GBMs. Increasing the expression of ASCL1 in PN CSCs results in suppression of self-renewal, promotion of differentiation and, most significantly, decrease in tumorigenesis, which is also reproduced by NDRG1 silencing. Conversely, both abrogation of ASCL1 expression in PN CSCs and enforcement of NDRG1 expression in either PN or MES CSCs induce proneural-to-mesenchymal transition (PMT) and enhanced mesenchymal features. Surprisingly, ASCL1 overexpression in MES CSCs increases malignant features and gives rise to a neuroendocrine-like secretory phenotype. Altogether, our results propose that the fine interplay between ASCL1 and its target NDRG1 might serve as potential subgroup-specific targetable vulnerability in GBM; enhancing ASCL1 expression in PN GBMs might reduce tumorigenesis, whereas repressing NDRG1 expression might be actionable to hamper the malignancy of GBM belonging to the MES subgroup.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Carcinogénesis/genética , Proteínas de Ciclo Celular/genética , Glioblastoma/genética , Péptidos y Proteínas de Señalización Intracelular/genética , Diferenciación Celular/genética , Línea Celular Tumoral , Proliferación Celular/genética , Autorrenovación de las Células/genética , Regulación Neoplásica de la Expresión Génica/genética , Glioblastoma/patología , Humanos , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Neuronas/metabolismo , Neuronas/patología , Transducción de SeñalRESUMEN
Impressões a partir da obrigação de ler a escrita de muitos alunos, viver a postergação de muitos e ter compilado alguns dizeres ferozes de renomados escritores, todos muito argutos em relação ao escrever. O texto combina fragmentos de autores com aforismos em torno da experiência que é escrever.
These text composed authors fragments with aphorims of a professor about the art of writing. Take off arguments by obligation to read the writing of many students, living their desperates and postpones around the experience that is to write. For that, compiled some fierce sayings from renowned writers, all very keen on writing.
Impresiones desde la obligación de leer los escritos de muchos alumnos, vivir la postergación de muchos y haber compilado algunos dichos feroces de renombrados escritores, todos muy astutos en relación al escribir. El texto combina fragmentos de autores con aforismos en torno a la experiéncia que es escribir.
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Lectura , Docentes , Comunicación Académica , Escritura Manual , Universidades , Acontecimientos que Cambian la VidaRESUMEN
Tuberous sclerosis complex (TSC) is a dominantly inherited disease caused by hyperactivation of the mTORC1 pathway and characterized by the development of hamartomas and benign tumors, including in the brain. Among the neurological manifestations associated with TSC, the tumor progression of static subependymal nodules (SENs) into subependymal giant cell astrocytomas (SEGAs) is one of the major causes of morbidity and shortened life expectancy. To date, mouse modeling has failed in reproducing these 2 lesions. Here we report that simultaneous hyperactivation of mTORC1 and Akt pathways by codeletion of Tsc1 and Pten, selectively in postnatal neural stem cells (pNSCs), is required for the formation of bona fide SENs and SEGAs. Notably, both lesions closely recapitulate the pathognomonic morphological and molecular features of the corresponding human abnormalities. The establishment of long-term expanding pNSC lines from mouse SENs and SEGAs made possible the identification of mTORC2 as one of the mediators conferring tumorigenic potential to SEGA pNSCs. Notably, in spite of concurrent Akt hyperactivation in mouse brain lesions, single mTOR inhibition by rapamycin was sufficient to strongly impair mouse SEGA growth. This study provides evidence that, concomitant with mTORC1 hyperactivation, sustained activation of Akt and mTORC2 in pNSCs is a mandatory step for the induction of SENs and SEGAs, and, at the same time, makes available an unprecedented NSC-based in vivo/in vitro model to be exploited for identifying actionable targets in TSC.
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Astrocitoma/metabolismo , Neoplasias Encefálicas/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Células-Madre Neurales/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Esclerosis Tuberosa/metabolismo , Animales , Astrocitoma/genética , Astrocitoma/patología , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Diana Mecanicista del Complejo 1 de la Rapamicina/genética , Ratones , Ratones Transgénicos , Células-Madre Neurales/patología , Proteínas Proto-Oncogénicas c-akt/genética , Esclerosis Tuberosa/genética , Esclerosis Tuberosa/patologíaRESUMEN
Precise control of the thyroid hormone (T3)-dependent transcriptional program is required by multiple cell systems, including muscle stem cells. Deciphering how this is achieved and how the T3 signal is controlled in stem cell niches is essentially unknown. We report that in response to proliferative stimuli such as acute skeletal muscle injury, type 3 deiodinase (D3), the thyroid hormone-inactivating enzyme, is induced in satellite cells where it reduces intracellular thyroid signaling. Satellite cell-specific genetic ablation of dio3 severely impairs skeletal muscle regeneration. This impairment is due to massive satellite cell apoptosis caused by exposure of activated satellite cells to the circulating TH. The execution of this proapoptotic program requires an intact FoxO3/MyoD axis, both genes positively regulated by intracellular TH. Thus, D3 is dynamically exploited in vivo to chronically attenuate TH signaling under basal conditions while also being available to acutely increase gene programs required for satellite cell lineage progression.
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Músculo Esquelético/citología , Células Madre/citología , Hormonas Tiroideas/metabolismo , Animales , Apoptosis/fisiología , Proliferación Celular/fisiología , Células Cultivadas , Proteína Forkhead Box O3 , Factores de Transcripción Forkhead/metabolismo , Inmunohistoquímica , Masculino , Ratones , Ratones Endogámicos C57BL , Células Satélite del Músculo Esquelético/citología , Células Satélite del Músculo Esquelético/metabolismo , Transducción de Señal/fisiologíaRESUMEN
Macrophages are one of the first barriers of host defence against pathogens. Beyond their role in innate immunity, macrophages play increasingly defined roles in orchestrating the healing of various injured tissues. Perturbations of macrophage function and/or activation may result in impaired regeneration and fibrosis deposition as described in several chronic pathological diseases. Heterogeneity and plasticity have been demonstrated to be hallmarks of macrophages. In response to environmental cues they display a proinflammatory (M1) or an alternative anti-inflammatory (M2) phenotype. A lot of evidence demonstrated that after acute injury M1 macrophages infiltrate early to promote the clearance of necrotic debris, whereas M2 macrophages appear later to sustain tissue healing. Whether the sequential presence of two different macrophage populations results from a dynamic shift in macrophage polarization or from the recruitment of new circulating monocytes is a subject of ongoing debate. In this paper, we discuss the current available information about the role that different phenotypes of macrophages plays after injury and during the remodelling phase in different tissue types, with particular attention to the skeletal muscle.
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Activación de Macrófagos/inmunología , Macrófagos/inmunología , Músculo Esquelético/inmunología , Músculo Esquelético/lesiones , Miositis/inmunología , Regeneración/inmunología , Cicatrización de Heridas/inmunología , Adaptación Fisiológica/inmunología , Animales , Humanos , Modelos Inmunológicos , Músculo Esquelético/patologíaRESUMEN
Given the extensive histomorphological heterogeneity of high-grade gliomas, in terms of extent of invasiveness, angiogenesis, and necrosis and the poor prognosis for patients despite the advancements made in therapeutic management. The identification of genes associated with these phenotypes will permit a better definition of glioma heterogeneity, which may ultimately lead to better treatment strategies. CXCR4, a cell surface chemokine receptor, is implicated in the growth, invasion, angiogenesis and metastasis in a wide range of malignant tumors, including gliomas. It is overexpressed in glioma cells according to tumor grade and in glioma tumor initiating cells. There have been various reports suggesting that CXCR4 is required for tumor proliferation, invasion, angiogenesis, and modulation of the immune response. It may also serve as a prognostic factor in characterizing subsets of glioblastoma multiforme, as patients with CXCR4-positive gliomas seem to have poorer prognosis after surgery. Aim of this review was to analyze the current literature on biological effects of CXCR4 activity and its role in glioma pathogenesis. A better understanding of CXCR4 pathway in glioma will lead to further investigation of CXCR4 as a novel putative therapeutic target.
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Neoplasias Encefálicas/fisiopatología , Glioma/fisiopatología , Receptores CXCR4/metabolismo , Animales , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/patología , Glioblastoma/tratamiento farmacológico , Glioblastoma/patología , Glioblastoma/fisiopatología , Glioma/tratamiento farmacológico , Glioma/patología , HumanosRESUMEN
The existence and identification of adult renal stem cells is a controversial issue. In this study, renal stem cells were identified from cultures of clonal human nephrospheres. The cultured nephrospheres exhibited the activation of stem cell pathways and contained cells at different levels of maturation. In each nephrosphere the presence of 1.12-1.25 cells mirroring stem cell properties was calculated. The nephrosphere cells were able to generate three-dimensional tubular structures in 3D cultures and in vivo. In clonal human nephrospheres a PKH(high) phenotype was isolated using PKH26 epifluorescence, which can identify quiescent cells within the nephrospheres. The PKH(high) cells, capable of self-renewal and of generating a differentiated epithelial, endothelial and podocytic progeny, can also survive in vivo maintaining the undifferentiated status. The PKH(high) status, together with a CD133(+)/CD24(-) phenotype, identified a homogeneous cell population displaying in vitro self-renewal and multipotency capacity. The resident adult renal stem cell population isolated from nephrospheres can be used for the study of mechanisms that regulate self-renewal and differentiation in adult renal tissue as well as in renal pathological conditions.
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Células Madre Adultas/citología , Riñón/citología , Compuestos Orgánicos/química , Antígeno AC133 , Células Madre Adultas/metabolismo , Células Madre Adultas/trasplante , Animales , Antígenos CD/metabolismo , Antígeno CD24/metabolismo , Técnicas de Cultivo de Célula , Diferenciación Celular , Células Cultivadas , Colorantes Fluorescentes/química , Glicoproteínas/metabolismo , Humanos , Ratones , Ratones Desnudos , Péptidos/metabolismo , Fenotipo , Trasplante HeterólogoRESUMEN
Inflammation plays a crucial role in muscle remodeling and repair after acute and chronic damage, in particular in muscular dystrophies, a heterogeneous group of genetic diseases leading to muscular degeneration. Defect of nitric oxide (NO) generation is a key pathogenic event in muscular dystrophies, thus NO donors have been explored as new therapeutics for this disease. We have investigated the immune-modulating effect of one of such drugs, molsidomine, able to slow the progression of muscular dystrophy in the α-Sarcoglican-null mice, a model for the limb girdle muscular dystrophy 2D, sharing several hallmarks of muscle degeneration with other muscular dystrophies. α-Sarcoglican-null mice were treated with molsidomine and drug effects on the inflammatory infiltrates and on muscle repair were assessed at selected time points. We found that molsidomine treatment modulates effectively the characteristics of the inflammatory infiltrate within dystrophic muscles, enhancing its healing function. Initially molsidomine amplified macrophage recruitment, promoting a more efficient clearance of cell debris and effective tissue regeneration. At a later stage molsidomine decreased significantly the extent of the inflammatory infiltrate, whose persistence exacerbates muscle damage: most of the remaining macrophages displayed characteristics of the transitional population, associated with reduced fibrosis and increased preservation of the muscle tissue. The dual action of molsidomine, the already known NO donation and the immunomodulatory function we now identified, suggests that it has a unique potential in tissue healing during chronic muscle damage. This, alongside its already approved use in human, makes molsidomine a drug with a significant therapeutic potential in muscular dystrophies.
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Inmunidad Innata/efectos de los fármacos , Molsidomina/metabolismo , Molsidomina/farmacología , Distrofias Musculares/inmunología , Óxido Nítrico/metabolismo , Animales , Modelos Animales de Enfermedad , Fibrosis , Humanos , Inflamación/inmunología , Inflamación/patología , Ratones , Distrofias Musculares/patología , FenotipoRESUMEN
By serving as the sole output of the cerebellar cortex, integrating a myriad of afferent stimuli, Purkinje cells (PCs) constitute the principal neuron in cerebellar circuits. Several neurodegenerative cerebellar ataxias feature a selective cell-autonomous loss of PCs, warranting the development of regenerative strategies. To date, very little is known as to the regulatory cascades controlling PC development. During central nervous system development, the proneural gene neurogenin 2 (Neurog2) contributes to many distinct neuronal types by specifying their fate and/or dictating development of their morphological features. By analyzing a mouse knock-in line expressing Cre recombinase under the control of Neurog2 cis-acting sequences we show that, in the cerebellar primordium, Neurog2 is expressed by cycling progenitors cell-autonomously fated to become PCs, even when transplanted heterochronically. During cerebellar development, Neurog2 is expressed in G1 phase by progenitors poised to exit the cell cycle. We demonstrate that, in the absence of Neurog2, both cell-cycle progression and neuronal output are significantly affected, leading to an overall reduction of the mature cerebellar volume. Although PC fate identity is correctly specified, the maturation of their dendritic arbor is severely affected in the absence of Neurog2, as null PCs develop stunted and poorly branched dendrites, a defect evident from the early stages of dendritogenesis. Thus, Neurog2 represents a key regulator of PC development and maturation.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/fisiología , Ciclo Celular , Cerebelo/crecimiento & desarrollo , Dendritas/fisiología , Proteínas del Tejido Nervioso/fisiología , Células de Purkinje/fisiología , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Linaje de la Célula , Cerebelo/fisiología , Femenino , Técnicas de Sustitución del Gen , Ratones , Ratones Endogámicos C57BL , Proteínas del Tejido Nervioso/genética , Neurogénesis/fisiología , Embarazo , Trasplante de Células Madre , Células Madre/fisiologíaRESUMEN
Congenital anomalies of the kidney and the urinary tract (CAKUT) represent a major source of morbidity and mortality in children. Several factors (PAX, SOX,WNT, RET, GDFN, and others) play critical roles during the differentiation process that leads to the formation of nephron epithelia. We have identified mutations in SOX17, an HMG-box transcription factor and Wnt signaling antagonist, in eight patients with CAKUT (seven vesico-ureteric reflux, one pelvic obstruction). One mutation, c.775T>A (p.Y259N), recurred in six patients. Four cases derived from two small families; renal scars with urinary infection represented the main symptom at presentation in all but two patients. Transfection studies indicated a 5-10-fold increase in the levels of the mutant protein relative to wild-type SOX17 in transfected kidney cells. Moreover we observed a corresponding increase in the ability of SOX17 p.Y259N to inhibit Wnt/ß-catenin transcriptional activity, which is known to regulate multiple stages of kidney and urinary tract development. In conclusion, SOX17 p.Y259N mutation is recurrent in patients with CAKUT. Our data shows that this mutation correlates with an inappropriate accumulation of SOX17-p.Y259N protein and inhibition of the ß-catenin/Wnt signaling pathway. These data indicate a role of SOX17 in human kidney and urinary tract development and implicate the SOX17-p.Y259N mutation as a causative factor in CAKUT.
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Anomalías Congénitas/genética , Riñón/anomalías , Mutación/genética , Factores de Transcripción SOXF/genética , Sistema Urinario/anomalías , Preescolar , Hibridación Genómica Comparativa , Familia , Femenino , Regulación de la Expresión Génica , Células HEK293 , Humanos , Hibridación Fluorescente in Situ , Masculino , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Linaje , Embarazo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factores de Transcripción SOXF/metabolismoRESUMEN
The embryonic cerebellum contains two germinative epithelia: the rhombic lip and the ventricular zone. While the lineage of glutamatergic neurons arising from the rhombic lip has been characterized, plenty remains to be learned about the factors giving rise to the array of ventricular zone-derived gamma-aminobutyric acid (GABA)ergic neurons. In the present study, we describe the expression of proneural genes Mash1/Ascl1, Ngn1/Neurog1, and Ngn2/Neurog2 in the cerebellar primordium at key stages of Purkinje cell and interneuron development, and compare them with the expression of other genes active in the same context. Our results indicate that Ngn1, Ngn2 and Mash1 are expressed at relevant stages of cerebellar neurogenesis in the prospective cerebellar nuclei and in the ventricular zone, excluding the Math1/Atoh1-positive rhombic lip. Their expression domains are only partially overlapping, suggesting that they may contribute selectively to ventricular zone regionalization, giving rise to the diversity of cerebellar GABA neurons and, possibly, Purkinje cell subtypes.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/biosíntesis , Cerebelo/embriología , Regulación del Desarrollo de la Expresión Génica , Proteínas del Tejido Nervioso/biosíntesis , Animales , Linaje de la Célula , Perfilación de la Expresión Génica , Hibridación in Situ , Ratones , Modelos Biológicos , Neuronas/metabolismo , Células de Purkinje/metabolismo , Factores de Tiempo , Distribución TisularRESUMEN
Myotubularin-related proteins (MTMRs) constitute a broad family of ubiquitously expressed phosphatases with 14 members in humans, of which eight are catalytically active phosphatases, while six are catalytically inactive. Active MTMRs possess 3-phosphatase activity toward both PtdIns3P and PtdIns(3, 5)P 2 poliphosphoinositides (PPIn), suggesting an involvement in intracellular trafficking and membrane homeostasis. Among MTMRs, catalytically active MTMR2 and inactive MTMR13 have a nonredundant function in nerve. Loss of either MTMR2 or MTMR13 causes Charcot-Marie-Tooth type 4B1 and B2 neuropathy, respectively, characterized by demyelination and redundant loops of myelin known as myelin outfoldings. In Mtmr2-null mouse nerves, these aberrant foldings occur at 3-4 weeks after birth, a time when myelination is established, and Schwann cells are still elongating to reach the final internodal length. Moreover, Mtmr2-specific ablation in Schwann cells is both sufficient and necessary to provoke CMT4B1 with myelin outfoldings. MTMR2 phospholipid phosphatase might regulate intracellular trafficking events and membrane homeostasis in Schwann cells during postnatal nerve development. In this review, we will discuss recent findings on the MTMR family with a major focus on MTMR2 and MTMR13 and their putative role in Schwann cell biology.
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Sistema Nervioso Periférico/enzimología , Isoformas de Proteínas/metabolismo , Proteínas Tirosina Fosfatasas no Receptoras/metabolismo , Células de Schwann/metabolismo , Animales , Enfermedad de Charcot-Marie-Tooth/clasificación , Enfermedad de Charcot-Marie-Tooth/metabolismo , Humanos , Vaina de Mielina/ultraestructura , Sistema Nervioso Periférico/anatomía & histología , Fosfatos de Fosfatidilinositol/metabolismo , Isoformas de Proteínas/genética , Proteínas Tirosina Fosfatasas no Receptoras/genética , Especificidad por SustratoRESUMEN
The RET gene is tightly regulated at the transcriptional level during embryo development, however in vitro experiments in cultured cells have failed to clarify the molecular mechanism of cell-type specificity of RET promoter activity. Therefore, we have generated transgenic mice in which the LacZ reporter gene is controlled by murine Ret promoter sequences to clarify in an in vivo model how this transcriptional regulation is achieved. We describe here the results of reporter gene expression in mice in which the transgene contained 380- and 1962-bp sequence upstream of the ATG start codon, derived from the mouse Ret promoter region, fused to the beta-galactosidase coding sequence. Transgenic mice showed well-defined patterns of beta-galactosidase staining obtained with both transgenes, suggesting that they were able per se to direct the reporter gene expression in specific districts, such as cranial ganglia, dorsal root ganglia, the heart and the kidney, partially recapitulating the profile of the endogenous Ret gene. In particular, proper expression in the developing excretory system seemed quite significant when considering that the appropriate regulation was obtained with a very short, 380 bp, fragment of Ret 5' flanking sequence.
Asunto(s)
Región de Flanqueo 5'/genética , Regulación del Desarrollo de la Expresión Génica/genética , Regiones Promotoras Genéticas/genética , Proteínas Proto-Oncogénicas c-ret/genética , Animales , Nervios Craneales/embriología , Nervios Craneales/metabolismo , Sistema Nervioso Entérico/embriología , Sistema Nervioso Entérico/metabolismo , Femenino , Corazón Fetal/embriología , Corazón Fetal/metabolismo , Ganglios/embriología , Ganglios/metabolismo , Ganglios Espinales/embriología , Ganglios Espinales/metabolismo , Inmunohistoquímica , Riñón/embriología , Riñón/metabolismo , Operón Lac/genética , Masculino , Ratones , Ratones Transgénicos , Páncreas/embriología , Páncreas/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , beta-Galactosidasa/genética , beta-Galactosidasa/metabolismoRESUMEN
The molecular characterization of balanced chromosomal rearrangements has often been a powerful tool for the positional identification of genes associated with specific diseases. In some instances, these rearrangements may be associated with a variety of different phenotypes, and thus establishing a genotype-phenotype correlation may be a complex process. However, molecular characterization of the rearrangement remains a useful tool for diagnoses or prognoses, or for identifying new genes and establishing a gene-to-function relationship. In this work we describe the characterization of a de novo balanced translocation t(2;6)(q24.3;q22.31) found in a patient with a complex phenotype. The major clinical finding was a severe neurological involvement. Thanks to the molecular characterization of this translocation we found that the rearrangement led to the truncation of the TCBA1 gene on chromosome 6q. We found that the gene is transcribed in different splice variants and is highly specific for the central nervous system. TCBA1 does not show any similarity with other known genes, and no information is available about its function. However, the gene appears to be well conserved among species, and we were able to infer the sequence of a putative mouse homolog of TCBA1. This allowed us to perform a more detailed expression study in mice, thus confirming its specificity for the nervous system. This finding is of particular interest because it suggests that TCBA1 may be correlated with the neurological phenotype of our patient, and possibly mutated in genetic diseases with a neurological phenotype.
Asunto(s)
Proteínas de la Membrana/genética , Enfermedades del Sistema Nervioso/genética , Eliminación de Secuencia , Translocación Genética , Empalme Alternativo , Animales , Secuencia de Bases , Encéfalo/metabolismo , Encéfalo/patología , Cromosomas Humanos Par 2 , Cromosomas Humanos Par 6 , Análisis Mutacional de ADN , Embrión de Mamíferos/metabolismo , Embrión de Mamíferos/patología , Humanos , Hibridación Fluorescente in Situ , Lactante , Cariotipificación , Masculino , Ratones , Datos de Secuencia Molecular , Enfermedades del Sistema Nervioso/diagnóstico , Enfermedades del Sistema Nervioso/patología , Fenotipo , ARN Mensajero/metabolismoRESUMEN
Regulation of the murine Ret gene has only been partially studied, specifically cis-acting promoter elements and related transcription factors, although in vivo experiments have shown the ability of Ret 5' flanking sequences to drive expression of transgenic reporter genes. This work describes the characterization of a 387-bp fragment of the Ret 5' flanking sequence with the ability to activate expression of the LacZ gene in transfected cells, in which SP1 recognition elements played a fundamental role. Using P19 teratocarcinoma cells differentiated by retinoic acid treatment, we observed an induction of Ret mRNA accompanied by a decrease of SP1 binding due to its proteolytic cleavage.