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Pancreatic cancer is a malignant tumor of the digestive system. It is highly aggressive, easily metastasizes, and extremely difficult to treat. This study aimed to analyze the genes that might regulate pancreatic cancer migration to provide an essential basis for the prognostic assessment of pancreatic cancer and individualized treatment. A CRISPR knockout library directed against 915 murine genes was transfected into TB 32047 cell line to screen which gene loss promoted cell migration. Next-generation sequencing and PinAPL.py- analysis was performed to identify candidate genes. We then assessed the effect of serine/threonine kinase 11 (STK11) knockout on pancreatic cancer by wound-healing assay, chick agnosia (CAM) assay, and orthotopic mouse pancreatic cancer model. We performed RNA sequence and Western blotting for mechanistic studies to identify and verify the pathways. After accelerated Transwell migration screening, STK11 was identified as one of the top candidate genes. Further experiments showed that targeted knockout of STK11 promoted the cell migration and increased liver metastasis in mice. Mechanistic analyses revealed that STK11 knockout influences blood vessel morphogenesis and is closely associated with the enhanced expression of phosphodiesterases (PDEs), especially PDE4D, PDE4B, and PDE10A. PDE4 inhibitor Roflumilast inhibited STK11-KO cell migration and tumor size, further demonstrating that PDEs are essential for STK11-deficient cell migration. Our findings support the adoption of therapeutic strategies, including Roflumilast, for patients with STK11-mutated pancreatic cancer in order to improve treatment efficacy and ultimately prolong survival.
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BACKGROUND AND AIMS: Inflammatory bowel diseases (IBD) are characterized by mucosal inflammation and sequential fibrosis formation, but the exact role of the hyperactive NLRP3 inflammasome in these processes is unclear. Thus, we studied the expression and function of the NLRP3 inflammasome in the context of inflammation and fibrosis in IBD. METHODS: We analysed intestinal NLRP3 expression in mucosal immune cells and fibroblasts from IBD patients and NLRP3-associated gene expression via single-cell RNA sequencing and microarray analyses. Furthermore, cytokine secretion of NLRP3 inhibitor treated blood and mucosal cells, as well as proliferation, collagen production, and cell death of NLRP3 inhibitor treated intestinal fibroblasts from IBD patients were studied. RESULTS: We found increased NLRP3 expression in the inflamed mucosa of IBD patients and NLRP3 inhibition led to reduced IL-1ß and IL-18 production in blood cells and diminished the bioactive form of mucosal IL-1ß. Single cell analysis identified overlapping expression patterns of NLRP3 and IL-1ß in classically activated intestinal macrophages and we also detected NLRP3 expression in CD163+ macrophages. In addition, NLRP3 expression was also found in intestinal fibroblasts from IBD patients. Inhibition of NLRP3 led to reduced proliferation of intestinal fibroblasts, which was associated with a marked decrease in production of collagen type I and type VI in IBD patients. Moreover, NLRP3 inhibition in intestinal fibroblasts induced autophagy, a cellular process involved in collagen degradation. CONCLUSIONS: In the presented study, we demonstrate that inhibiting NLRP3 might pave the way for novel therapeutic approaches in IBD, especially to prevent the severe complication of intestinal fibrosis formation.
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Enfermedades Inflamatorias del Intestino , Proteína con Dominio Pirina 3 de la Familia NLR , Humanos , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Inflamasomas/metabolismo , Membrana Mucosa/metabolismo , Interleucina-1beta/metabolismo , Inflamación , Fibroblastos/metabolismo , Colágeno , FibrosisRESUMEN
Lymphangiogenesis is a key player in several diseases such as tumor metastasis, obesity, and graft rejection. Endogenous regulation of lymphangiogenesis is only partly understood. Here we use the normally avascular cornea as a model to identify endogenous regulators of lymphangiogenesis. Quantitative trait locus analysis of a large low-lymphangiogenic BALB/cN x high-lymphangiogenic C57BL/6 N intercross and prioritization by whole-transcriptome sequencing identify a novel gene responsible for differences in lymphatic vessel architecture on chromosome 17, the cystathionine ß-synthase (Cbs). Inhibition of CBS in lymphatic endothelial cells results in reduce proliferation, migration, altered tube-formation, and decrease expression of vascular endothelial growth factor (VEGF) receptor 2 (VEGF-R2) and VEGF-R3, but not their ligands VEGF-C and VEGF-D. Also in vivo inflammation-induced lymphangiogenesis is significantly reduce in C57BL/6 N mice after pharmacological inhibition of CBS. The results confirm CBS as a novel endogenous regulator of lymphangiogenesis acting via VEGF receptor 2 and 3-regulation and open new treatment avenues in diseases associated with pathologic lymphangiogenesis.
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Cistationina betasintasa , Linfangiogénesis , Animales , Cistationina betasintasa/genética , Cistationina betasintasa/metabolismo , Células Endoteliales/metabolismo , Ratones , Ratones Endogámicos C57BL , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Under normal conditions, the cornea, being the transparent "windscreen" of the eye, is free of both blood and lymphatic vessels. However, various diseases of the eye, like infections, can interfere with the balance between promoting and inhibiting factors, which leads to ingrowth of blood and lymphatic vessels. The newly formed lymphatic vessels increase the risk of graft rejection after subsequent corneal transplantation. Corneal transplantation is one of the most commonly performed transplantations worldwide, with more than 40,000 surgeries per year in Europe. To date, various anti-hem- and anti-lymphangiogenic treatment strategies have been developed specifically for the corneal vascular endothelial growth factor (VEGF) pathway. Currently, however, no treatment strategies are clinically available to specifically modulate lymphangiogenesis. In this review, we will give an overview about endogenous regulators of hem- and lymphangiogenesis and discuss potential new strategies for targeting pathological lymphangiogenesis. Furthermore, we will review recently identified modulators and demonstrate that the cornea is a suitable model for the identification of novel endogenous modulators of lymphangiogenesis. The identification of novel modulators of lymphangiogenesis and a better understanding of the signaling pathways involved will contribute to the development of new therapeutic targets for the treatment of pathological lymphangiogenesis. This, in turn, will improve graft rejection, not only for the cornea.
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OBJECTIVE: Cancer-associated fibroblasts (CAFs) influence the tumour microenvironment and tumour growth. However, the role of CAFs in colorectal cancer (CRC) development is incompletely understood. DESIGN: We quantified phosphorylation of STAT3 (pSTAT3) expression in CAFs of human colon cancer tissue using a tissue microarray (TMA) of 375 patients, immunofluorescence staining and digital pathology. To investigate the functional role of CAFs in CRC, we took advantage of two murine models of colorectal neoplasia and advanced imaging technologies. In loss-of-function and gain-of-function experiments, using genetically modified mice with collagen type VI (COLVI)-specific signal transducer and activator of transcription 3 (STAT3) targeting, we evaluated STAT3 signalling in fibroblasts during colorectal tumour development. We performed a comparative gene expression profiling by whole genome RNA-sequencing of fibroblast subpopulations (COLVI+ vs COLVI-) on STAT3 activation (IL-6 vs IL-11). RESULTS: The analysis of pSTAT3 expression in CAFs of human TMAs revealed a negative correlation of increased stromal pSTAT3 expression with the survival of colon cancer patients. In the loss-of-function and gain-of-function approach, we found a critical role of STAT3 activation in fibroblasts in driving colorectal tumourigenesis in vivo. With different imaging technologies, we detected an expansion of activated fibroblasts in colorectal neoplasias. Comparative gene expression profiling of fibroblast subpopulations on STAT3 activation revealed the regulation of transcriptional patterns associated with angiogenesis. Finally, the blockade of proangiogenic signalling significantly reduced colorectal tumour growth in mice with constitutive STAT3 activation in COLVI+ fibroblasts. CONCLUSION: Altogether our work demonstrates a critical role of STAT3 activation in CAFs in CRC development.
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Neoplasias Colorrectales/etiología , Interleucina-11/metabolismo , Interleucina-6/metabolismo , Factor de Transcripción STAT3/metabolismo , Animales , Colon/metabolismo , Neoplasias Colorrectales/diagnóstico , Fibroblastos/metabolismo , Humanos , Ratones , Fosforilación , Pronóstico , Análisis de Matrices Tisulares , TranscriptomaRESUMEN
Height is a heritable and highly heterogeneous trait. Short stature affects 3% of the population and in most cases is genetic in origin. After excluding known causes, 67% of affected individuals remain without diagnosis. To identify novel candidate genes for short stature, we performed exome sequencing in 254 unrelated families with short stature of unknown cause and identified variants in 63 candidate genes in 92 (36%) independent families. Based on systematic characterization of variants and functional analysis including expression in chondrocytes, we classified 13 genes as strong candidates. Whereas variants in at least two families were detected for all 13 candidates, two genes had variants in 6 (UBR4) and 8 (LAMA5) families, respectively. To facilitate their characterization, we established a clustered network of 1025 known growth and short stature genes, which yielded 29 significantly enriched clusters, including skeletal system development, appendage development, metabolic processes, and ciliopathy. Eleven of the candidate genes mapped to 21 of these clusters, including CPZ, EDEM3, FBRS, IFT81, KCND1, PLXNA3, RASA3, SLC7A8, UBR4, USP45, and ZFHX3. Fifty additional growth-related candidates we identified await confirmation in other affected families. Our study identifies Mendelian forms of growth retardation as an important component of idiopathic short stature.
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Enanismo/genética , Exoma , Herencia Multifactorial , Niño , Preescolar , Proteínas del Citoesqueleto/genética , Femenino , Humanos , Lactante , Masculino , Secuenciación del ExomaRESUMEN
Immune sensing of Mycobacterium tuberculosis relies on recognition by macrophages. Mycobacterial cord factor, trehalose-6,6'-dimycolate (TDM), is the most abundant cell wall glycolipid and binds to the C-type lectin receptor (CLR) MINCLE. To explore the kinase signaling linking the TDM-MINCLE interaction to gene expression, we employed quantitative phosphoproteome analysis. TDM caused upregulation of 6.7% and suppressed 3.8% of the 14,000 phospho-sites identified on 3727 proteins. MINCLE-dependent phosphorylation was observed for canonical players of CLR signaling (e.g. PLCγ, PKCδ), and was enriched for PKCδ and GSK3 kinase motifs. MINCLE-dependent activation of the PI3K-AKT-GSK3 pathway contributed to inflammatory gene expression and required the PI3K regulatory subunit p85α. Unexpectedly, a substantial fraction of TDM-induced phosphorylation was MINCLE-independent, a finding paralleled by transcriptome data. Bioinformatics analysis of both data sets concurred in the requirement for MINCLE for innate immune response pathways and processes. In contrast, MINCLE-independent phosphorylation and transcriptome responses were linked to cell cycle regulation. Collectively, our global analyses show substantial reprogramming of macrophages by TDM and reveal a dichotomy of MINCLE-dependent and -independent signaling linked to distinct biological responses.
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Factores Cordón/metabolismo , Lectinas Tipo C/metabolismo , Macrófagos/metabolismo , Proteínas de la Membrana/metabolismo , Fosfoproteínas/metabolismo , Proteoma/metabolismo , Proteómica , Transducción de Señal , Animales , Ciclo Celular/genética , Proliferación Celular/genética , Supervivencia Celular/genética , Factores Cordón/farmacología , Citocinas/metabolismo , Activación Enzimática/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Glucolípidos/metabolismo , Cinética , Activación de Macrófagos/efectos de los fármacos , Activación de Macrófagos/genética , Macrófagos/efectos de los fármacos , Ratones Endogámicos C57BL , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Mycobacterium tuberculosis/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Fosforilación/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal/efectos de los fármacos , Quinasa Syk/metabolismo , Transcriptoma/genética , Trehalosa/metabolismoRESUMEN
Lymphangiogenesis is critically involved in tissue fluid balance, graft rejection, and tumor metastasis. Endogenous regulation of lymphangiogenesis is poorly understood. Herein, we use the lymphatic vessel architecture at the limbal border of the normally avascular cornea, a quantitative trait under strong genetic influence, as a model system to identify new candidate genes regulating lymphangiogenesis. Comparing low-lymphangiogenic BALB/cN with high-lymphangiogenic C57BL/6N mice, we performed quantitative trait loci analysis of five phenotypes in a large BALB/cN × C57BL/6N intercross (n = 795) and identified three to eight genome-wide significant loci, the strongest on chromosome 7 containing tyrosinase (Tyr). Tyrosinase-negative mice showed significantly increased limbal lymph vascularized areas, a higher number of lymphatic vessel end points, and branching points and increased inflammation-induced lymphangiogenesis. These findings confirm that tyrosinase is a novel lymphangiogenesis regulator in developmental and inflammatory lymphangiogenesis. Our findings link melanin synthesis with lymphangiogenesis and open new treatment options in lymphangiogenesis-related diseases.
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Linfangiogénesis , Vasos Linfáticos/enzimología , Melaninas/biosíntesis , Monofenol Monooxigenasa/metabolismo , Animales , Cromosomas de los Mamíferos/genética , Cromosomas de los Mamíferos/metabolismo , Femenino , Sitios Genéticos , Vasos Linfáticos/patología , Masculino , Melaninas/genética , Ratones , Ratones Transgénicos , Monofenol Monooxigenasa/genéticaRESUMEN
PurposeShort stature is a common condition of great concern to patients and their families. Mostly genetic in origin, the underlying cause often remains elusive due to clinical and genetic heterogeneity.MethodsWe systematically phenotyped 565 patients where common nongenetic causes of short stature were excluded, selected 200 representative patients for whole-exome sequencing, and analyzed the identified variants for pathogenicity and the affected genes regarding their functional relevance for growth.ResultsBy standard targeted diagnostic and phenotype assessment, we identified a known disease cause in only 13.6% of the 565 patients. Whole-exome sequencing in 200 patients identified additional mutations in known short-stature genes in 16.5% of these patients who manifested only part of the symptomatology. In 15.5% of the 200 patients our findings were of significant clinical relevance. Heterozygous carriers of recessive skeletal dysplasia alleles represented 3.5% of the cases.ConclusionA combined approach of systematic phenotyping, targeted genetic testing, and whole-exome sequencing allows the identification of the underlying cause of short stature in at least 33% of cases, enabling physicians to improve diagnosis, treatment, and genetic counseling. Exome sequencing significantly increases the diagnostic yield and consequently care in patients with short stature.
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Estatura/genética , Femenino , Pruebas Genéticas , Heterocigoto , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Masculino , Mutación , Linaje , Fenotipo , Análisis de Secuencia de ADN/métodos , Secuenciación del Exoma/métodosRESUMEN
Variants affecting the function of different subunits of the BAF chromatin-remodelling complex lead to various neurodevelopmental syndromes, including Coffin-Siris syndrome. Furthermore, variants in proteins containing PHD fingers, motifs recognizing specific histone tail modifications, have been associated with several neurological and developmental-delay disorders. Here, we report eight heterozygous de novo variants (one frameshift, two splice site, and five missense) in the gene encoding the BAF complex subunit double plant homeodomain finger 2 (DPF2). Affected individuals share common clinical features described in individuals with Coffin-Siris syndrome, including coarse facial features, global developmental delay, intellectual disability, speech impairment, and hypoplasia of fingernails and toenails. All variants occur within the highly conserved PHD1 and PHD2 motifs. Moreover, missense variants are situated close to zinc binding sites and are predicted to disrupt these sites. Pull-down assays of recombinant proteins and histone peptides revealed that a subset of the identified missense variants abolish or impaire DPF2 binding to unmodified and modified H3 histone tails. These results suggest an impairment of PHD finger structural integrity and cohesion and most likely an aberrant recognition of histone modifications. Furthermore, the overexpression of these variants in HEK293 and COS7 cell lines was associated with the formation of nuclear aggregates and the recruitment of both wild-type DPF2 and BRG1 to these aggregates. Expression analysis of truncating variants found in the affected individuals indicated that the aberrant transcripts escape nonsense-mediated decay. Altogether, we provide compelling evidence that de novo variants in DPF2 cause Coffin-Siris syndrome and propose a dominant-negative mechanism of pathogenicity.
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Anomalías Múltiples/genética , Proteínas de Unión al ADN/genética , Cara/anomalías , Deformidades Congénitas de la Mano/genética , Discapacidad Intelectual/genética , Micrognatismo/genética , Mutación/genética , Cuello/anomalías , Subunidades de Proteína/genética , Adolescente , Secuencia de Aminoácidos , Animales , Células COS , Niño , Preescolar , Chlorocebus aethiops , Proteínas de Unión al ADN/química , Facies , Femenino , Células HEK293 , Histonas/metabolismo , Humanos , Masculino , Fenotipo , Factores de TranscripciónRESUMEN
Alcohol use disorders are major psychiatric disorders. Correlational studies in humans suggested organizational hormonal effects during embryonic development as a risk factor for adult alcohol dependence. Permanent changes can be induced by the activity of sex hormones, like testosterone. Here, we demonstrate a relationship between prenatal androgen receptor (AR)-activation and adult alcohol as well as water drinking in mice in a sex-dependent fashion. Prenatal AR inhibition using the antagonist flutamide decreased adult male alcohol consumption. In contrast, prenatal AR activation by dihydrotestosterone (DHT) led to an increase in adult alcohol consumption in females. These effects were different in adult water drinking, flutamide increased water consumption in females and DHT increased water consumption in males. Prenatal flutamide reduced locomotion and anxiety in adult males but was ineffective in females. We found that prenatal AR activation controls adult levels of monoaminergic modulatory transmitters in the brain and blood hormone levels in a sex-specific way. RNA-Seq analysis confirmed a prenatal AR mediated control of adult expression of alcohol drinking-related genes like Bdnf and Per2. These findings demonstrate that prenatal androgen activity is a risk factor for the establishment of alcohol consumption in adults by its organizational effects.
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Consumo de Bebidas Alcohólicas , Antagonistas de Receptores Androgénicos/farmacología , Andrógenos/farmacología , Dihidrotestosterona/farmacología , Conducta de Ingestión de Líquido/efectos de los fármacos , Flutamida/farmacología , Efectos Tardíos de la Exposición Prenatal/metabolismo , Receptores Androgénicos/metabolismo , Animales , Factor Neurotrófico Derivado del Encéfalo/efectos de los fármacos , Factor Neurotrófico Derivado del Encéfalo/genética , Conducta de Ingestión de Líquido/fisiología , Femenino , Expresión Génica/efectos de los fármacos , Masculino , Ratones , Proteínas Circadianas Period/efectos de los fármacos , Proteínas Circadianas Period/genética , Embarazo , ARN Mensajero/efectos de los fármacos , ARN Mensajero/metabolismo , Factores Sexuales , AguaRESUMEN
Eleven-nineteen leukemia (ENL) is a chromatin reader present in complexes stimulating transcriptional elongation. It is fused to mixed-lineage leukemia (MLL) in leukemia, and missense mutations have been identified in Wilms tumor and acute myeloid leukemia. Here we demonstrate that ENL overcomes polycomb silencing through recruitment of PAF1 via the conserved YEATS domain, which recognizes acetylated histone H3. PAF1 was responsible for antirepressive activities of ENL in vitro, and it determined the transforming potential of MLL-ENL. MLL-ENL target loci showed supraphysiological PAF1 binding, hyperubiquitination of histone H2B and hypomodification with H2AUb, resulting in accelerated transcription rates. YEATS mutations induced a gain of function, transforming primary hematopoietic cells in vitro and in transplantation assays through aberrant transcription and H2B ubiquitination of Hoxa9 and Meis1 Mechanistically, H3 and PAF1 competed for ENL interaction, with activating mutations favoring PAF1 binding, whereas the MLL moiety provided a constitutive PAF1 tether allowing MLL fusions to circumvent H3 competition.
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Proteínas Portadoras/metabolismo , Transformación Celular Neoplásica , Proteínas de Unión al ADN/metabolismo , Silenciador del Gen , Leucemia/genética , Proteínas del Grupo Polycomb/genética , Factores de Transcripción/metabolismo , Animales , Proteínas Portadoras/química , Proteínas Portadoras/genética , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/metabolismo , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/genética , Regulación Leucémica de la Expresión Génica , Células HEK293 , Histonas/metabolismo , Humanos , Leucemia/metabolismo , Leucemia/patología , Ratones , Ratones Endogámicos BALB C , Mutación , Unión Proteica/genética , Dominios Proteicos/genética , Procesamiento Proteico-Postraduccional , Factores de Transcripción/química , Factores de Transcripción/genéticaRESUMEN
Short stature is a common pediatric disorder affecting 3% of the population. However, the clinical variability and genetic heterogeneity prevents the identification of the underlying cause in about 80% of the patients. Recently, heterozygous mutations in the ACAN gene coding for the proteoglycan aggrecan, a main component of the cartilage matrix, were associated with idiopathic short stature. To ascertain the prevalence of ACAN mutations and broaden the phenotypic spectrum in patients with idiopathic short stature we performed sequence analyses in 428 families. We identified heterozygous nonsense mutations in four and potentially disease-causing missense variants in two families (1.4%). These patients presented with a mean of -3.2 SDS and some suggestive clinical characteristics. The results suggest heterozygous mutations in ACAN as a common cause of isolated as well as inherited idiopathic short stature.
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Agrecanos/genética , Estatura/genética , Trastornos del Crecimiento/genética , Fenotipo , Niño , Análisis Mutacional de ADN , Femenino , Pruebas Genéticas , Heterocigoto , Humanos , Masculino , Mutación , LinajeRESUMEN
BACKGROUND & AIMS: We investigated the roles of interleukin 28A (also called IL28A or interferon λ2) in intestinal epithelial cell (IEC) activation, studying its effects in mouse models of inflammatory bowel diseases (IBD) and intestinal mucosal healing. METHODS: Colitis was induced in C57BL/6JCrl mice (controls), mice with IEC-specific disruption of Stat1 (Stat1IEC-KO), mice with disruption of the interferon λ receptor 1 gene (Il28ra-/-), and mice with disruption of the interferon regulatory factor 3 gene (Irf3-/-), with or without disruption of Irf7 (Irf7-/-). We used high-resolution mini-endoscopy and in vivo imaging methods to assess colitis progression. We used 3-dimensional small intestine and colon organoids, along with RNA-Seq and gene ontology methods, to characterize the effects of IL28 on primary IECs. We studied the effects of IL28 on the human intestinal cancer cell line Caco-2 in a wound-healing assay, and in mice colon wounds. Colonic biopsies and resected tissue from patients with IBD (n = 62) and patients without colon inflammation (controls, n = 23) were analyzed by quantitative polymerase chain rection to measure expression of IL28A, IL28RA, and other related cytokines; biopsy samples were also analyzed by immunofluorescence to identify sources of IL28 production. IECs were isolated from patient tissues and incubated with IL28; signal transducer and activator of transcription 1 (STAT1) phosphorylation was measured by immunoblots and confocal imaging. RESULTS: Lamina propria cells in colon tissues of patients with IBD, and mice with colitis, had increased expression of IL28 compared with controls; levels of IL28R were increased in the colonic epithelium of patients with IBD and mice with colitis. Administration of IL28 induced phosphorylation of STAT1 in primary human and mouse IECs, increasing with dose. Il28ra-/-, Irf3-/-, Irf3-/-Irf7-/-, as well as Stat1IEC-KO mice, developed more severe colitis after administration of dextran sulfate sodium than control mice, with reduced epithelial restitution. Il28ra-/- and Stat1IEC-KO mice also developed more severe colitis in response to oxazolone than control mice. We found IL28 to induce phosphorylation (activation) of STAT1 in epithelial cells, leading to their proliferation in organoid culture. Administration of IL28 to mice with induced colonic wounds promoted mucosal healing. CONCLUSIONS: IL28 controls proliferation of IECs in mice with colitis and accelerates mucosal healing by activating STAT1. IL28 might be developed as a therapeutic agent for patients with IBD.
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Colitis/metabolismo , Enfermedades Inflamatorias del Intestino/metabolismo , Interleucinas/metabolismo , Factor de Transcripción STAT1/metabolismo , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Animales , Células CACO-2 , Proliferación Celular , Colitis/inducido químicamente , Colitis/genética , Colitis/patología , Células Dendríticas/metabolismo , Sulfato de Dextran , Modelos Animales de Enfermedad , Células Epiteliales , Femenino , Expresión Génica , Humanos , Enfermedades Inflamatorias del Intestino/genética , Factor 3 Regulador del Interferón/genética , Factor 7 Regulador del Interferón/genética , Interleucinas/genética , Interleucinas/farmacología , Mucosa Intestinal/metabolismo , Masculino , Ratones , Ratones Noqueados , Persona de Mediana Edad , Organoides , Fosforilación , Receptores de Citocinas/genética , Receptores de Citocinas/metabolismo , Receptores de Interferón/genética , Factor de Transcripción STAT1/genética , Transducción de Señal , Cicatrización de Heridas , Adulto JovenRESUMEN
Patients with the Mayer-Rokitansky-Küster-Hauser syndrome (MRKH) have a congenital utero-vaginal cervical aplasia, but normal or hypoplastic adnexa and develop with normal female phenotype. Some reports mostly demonstrated regular steroid hormone levels in small MRKH cohorts including single MRKH patients with hyperandrogenemia and a clinical presentationof hirsutism and acne has also been shown. Genetically a correlation of WNT4 mutations with singular MRKH patients and hyperandrogenemia was noted. This study analyzed the hormone status of 215 MRKH patients by determining the levels of luteinizing hormone (LH), follicle-stimulating hormone (FSH), estradiol, 17-OH progesterone, testosterone, dehydroepiandrosterone sulfate (DHEAS), sex hormone-binding globulin (SHBG) and prolactin to determine the incidence of hyperandrogenemia and hyperprolactinemia in MRKH patients. Additional calculations and a ratio of free androgen index and biologically active testosterone revealed a hyperandrogenemia rate of 48.3%, hyperprolactinemia of 9.8% and combined hyperandrogenemia and hyperprolactinemia of 4.2% in MRKH patients. The rates of hirsutism, acne and especially polycystic ovary syndrome (PCOS) were in the normal range of the population and showed no correlation with hyperandrogenemia. A weekly hormone assessment over 30 days comparing 5 controls and 7 MRKH patients revealed high androgen and prolactin, but lower LH/FSH and SHBG levels with MRKH patients. The sequencing of WNT4, WNT5A, WNT7A and WNT9B demonstrated no significant mutations correlating with hyperandrogenemia. Taken together, this study shows that over 52% of MRKH patients have hyperandrogenemia without clinical presentation and 14% hyperprolactinemia, which appeals for general hormone assessment and adjustments of MRKH patients.
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Anomalías Congénitas/fisiopatología , Hiperandrogenismo/etiología , Hiperprolactinemia/etiología , Anomalías Urogenitales/complicaciones , Útero/anomalías , Vagina/anomalías , Adulto , Femenino , Humanos , Hiperandrogenismo/diagnóstico , Hiperprolactinemia/diagnóstico , Pronóstico , SíndromeRESUMEN
Alcohol is a widely consumed drug that can lead to addiction and severe brain damage. However, alcohol is also used as self-medication for psychiatric problems, such as depression, frequently resulting in depression-alcoholism comorbidity. Here, we identify the first molecular mechanism for alcohol use with the goal to self-medicate and ameliorate the behavioral symptoms of a genetically induced innate depression. An induced over-expression of acid sphingomyelinase (ASM), as was observed in depressed patients, enhanced the consumption of alcohol in a mouse model of depression. ASM hyperactivity facilitates the establishment of the conditioned behavioral effects of alcohol, and thus drug memories. Opposite effects on drinking and alcohol reward learning were observed in animals with reduced ASM function. Importantly, free-choice alcohol drinking-but not forced alcohol exposure-reduces depression-like behavior selectively in depressed animals through the normalization of brain ASM activity. No such effects were observed in normal mice. ASM hyperactivity caused sphingolipid and subsequent monoamine transmitter hypo-activity in the brain. Free-choice alcohol drinking restores nucleus accumbens sphingolipid- and monoamine homeostasis selectively in depressed mice. A gene expression analysis suggested strong control of ASM on the expression of genes related to the regulation of pH, ion transmembrane transport, behavioral fear response, neuroprotection and neuropeptide signaling pathways. These findings suggest that the paradoxical antidepressant effects of alcohol in depressed organisms are mediated by ASM and its control of sphingolipid homeostasis. Both emerge as a new treatment target specifically for depression-induced alcoholism.
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Antidepresivos/uso terapéutico , Depresión/tratamiento farmacológico , Etanol/uso terapéutico , Homeostasis/genética , Esfingolípidos/metabolismo , Esfingomielina Fosfodiesterasa/metabolismo , Animales , Conducta de Elección/efectos de los fármacos , Condicionamiento Operante/efectos de los fármacos , Depresión/genética , Etanol/sangre , Preferencias Alimentarias/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/genética , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Reflejo de Enderezamiento/efectos de los fármacos , Reflejo de Enderezamiento/genética , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Esfingomielina Fosfodiesterasa/genética , Superóxido Dismutasa/genética , Superóxido Dismutasa/metabolismoRESUMEN
Williams-Beuren syndrome (WBS) is a relatively common, clinically recognizable microdeletion syndrome. In most cases the typical heterozygous deletion of 1.5 Mb on chromosome 7q11.23 spanning about 26 genes can be identified. Also some larger or smaller atypical deletions have been reported and associated with additional or atypical phenotypic aspects. We report on an individual with typical WBS due to the common deletion and with refractory infantile spasms. Using trio-exome sequencing, we identified a de novo truncating variant c.1200del, p (Lys401Serfs*25) in GABRA1 as the likely cause of the early onset epilepsy. This unique case not only allows to further define the phenotypic spectrum of infantile epileptic encephalopathy associated with rare de novo GABRA1 variants but exemplifies the need for a sensitive review of unclear associations in clinically defined syndromes and for extended diagnostic work-up in individuals with unusual presentations of a genetically confirmed diagnosis.
Asunto(s)
Epilepsia/genética , Receptores de GABA-A/genética , Espasmos Infantiles/genética , Síndrome de Williams/genética , Epilepsia/complicaciones , Epilepsia/fisiopatología , Exoma/genética , Humanos , Lactante , Recién Nacido , Masculino , Espasmos Infantiles/complicaciones , Espasmos Infantiles/fisiopatología , Síndrome de Williams/fisiopatologíaRESUMEN
Multiple system atrophy (MSA) is a rare atypical parkinsonian disorder characterized by a rapidly progressing clinical course and at present without any efficient therapy. Neuropathologically, myelin loss and neurodegeneration are associated with α-synuclein accumulation in oligodendrocytes, but underlying pathomechanisms are poorly understood. Here, we analyzed the impact of oligodendrocytic α-synuclein on the formation of myelin sheaths to define a potential interventional target for MSA. Post-mortem analyses of MSA patients and controls were performed to quantify myelin and oligodendrocyte numbers. As pre-clinical models, we used transgenic MSA mice, a myelinating stem cell-derived oligodendrocyte-neuron co-culture, and primary oligodendrocytes to determine functional consequences of oligodendrocytic α-synuclein overexpression on myelination. We detected myelin loss accompanied by preserved or even increased numbers of oligodendrocytes in post-mortem MSA brains or transgenic mouse forebrains, respectively, indicating an oligodendrocytic dysfunction in myelin formation. Corroborating this observation, overexpression of α-synuclein in primary and stem cell-derived oligodendrocytes severely impaired myelin formation, defining a novel α-synuclein-linked pathomechanism in MSA. We used the pro-myelinating activity of the muscarinic acetylcholine receptor antagonist benztropine to analyze the reversibility of the myelination deficit. Transcriptome profiling of primary pre-myelinating oligodendrocytes demonstrated that benztropine readjusts myelination-related processes such as cholesterol and membrane biogenesis, being compromised by oligodendrocytic α-synuclein. Additionally, benztropine restored the α-synuclein-induced myelination deficit of stem cell-derived oligodendrocytes. Strikingly, benztropine also ameliorated the myelin deficit in transgenic MSA mice, resulting in a prevention of neuronal cell loss. In conclusion, this study defines the α-synuclein-induced myelination deficit as a novel and crucial pathomechanism in MSA. Importantly, the reversible nature of this oligodendrocytic dysfunction opens a novel avenue for an intervention in MSA.
Asunto(s)
Antiparkinsonianos/farmacología , Benzotropina/farmacología , Atrofia de Múltiples Sistemas/tratamiento farmacológico , Atrofia de Múltiples Sistemas/metabolismo , alfa-Sinucleína/metabolismo , Animales , Encéfalo/diagnóstico por imagen , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Encéfalo/patología , Muerte Celular/efectos de los fármacos , Células Cultivadas , Técnicas de Cocultivo , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Gliosis/metabolismo , Gliosis/patología , Gliosis/prevención & control , Masculino , Ratones Transgénicos , Atrofia de Múltiples Sistemas/diagnóstico por imagen , Atrofia de Múltiples Sistemas/patología , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Neuronas/patología , Oligodendroglía/efectos de los fármacos , Oligodendroglía/metabolismo , Oligodendroglía/patología , Ratas Wistar , Células Madre/efectos de los fármacos , Células Madre/metabolismo , Células Madre/patología , Transcriptoma/efectos de los fármacos , alfa-Sinucleína/genéticaRESUMEN
MLL fusions are leukemogenic transcription factors that enhance transcriptional elongation through modification of chromatin and RNA Pol II. Global transcription rates and chromatin changes accompanying the transformation process induced by MLL-ENL were monitored by nascent RNA-seq and ChIP-seq, revealing 165 direct target genes separated into two distinct clades. ME5 genes bound MLL-ENL at the promoter, relied on DOT1L-mediated histone methylation, and coded preferentially for transcription factors, including many homeobox genes. A distinct ME3 group accumulated MLL-ENL beyond the termination site, was dependent on P-TEFb-mediated phosphorylation of RNA Pol II for transcription, and translated mainly into proteins involved in RNA biology and ribosome assembly. This dichotomy was reflected by a differential sensitivity toward small molecule inhibitors, suggesting the possibility of a combinatorial strategy for treatment of MLL-induced leukemia.