RESUMEN
BACKGROUND: The VPS50 protein functions in synaptic and dense core vesicle acidification, and perturbations of VPS50 function produce behavioral changes in Caenorhabditis elegans. Patients with mutations in VPS50 show severe developmental delay and intellectual disability, characteristics that have been associated with autism spectrum disorders (ASDs). The mechanisms that link VPS50 mutations to ASD are unknown. RESULTS: To examine the role of VPS50 in mammalian brain function and behavior, we used the CRISPR/Cas9 system to generate knockouts of VPS50 in both cultured murine cortical neurons and living mice. In cultured neurons, KO of VPS50 did not affect the number of synaptic vesicles but did cause mislocalization of the V-ATPase V1 domain pump and impaired synaptic activity, likely as a consequence of defects in vesicle acidification and vesicle content. In mice, mosaic KO of VPS50 in the hippocampus altered synaptic transmission and plasticity and generated robust cognitive impairments. CONCLUSIONS: We propose that VPS50 functions as an accessory protein to aid the recruitment of the V-ATPase V1 domain to synaptic vesicles and in that way plays a crucial role in controlling synaptic vesicle acidification. Understanding the mechanisms controlling behaviors and synaptic function in ASD-associated mutations is pivotal for the development of targeted interventions, which may open new avenues for therapeutic strategies aimed at ASD and related conditions.
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Ratones Noqueados , Vesículas Sinápticas , Animales , Ratones , Conducta Animal/fisiología , Encéfalo/metabolismo , Neuronas/metabolismo , Neuronas/fisiología , Sinapsis/metabolismo , Sinapsis/fisiología , Transmisión Sináptica , Vesículas Sinápticas/metabolismo , ATPasas de Translocación de Protón Vacuolares/metabolismo , ATPasas de Translocación de Protón Vacuolares/genética , Proteínas de Transporte Vesicular/genética , Proteínas de Transporte Vesicular/metabolismoRESUMEN
Depressive disorders (DD) have affected millions of people worldwide. Venlafaxine, antidepressant of the class of serotonin and norepinephrine reuptake inhibitors, has been prescribed for the treatment of DD. In rat testes, venlafaxine induces testosterone (T) aromatization and increases estrogen levels. Aromatase is a key enzyme for the formation of estrogen in the epididymis, an essential organ for male fertility. We investigated the impact of serotonergic/noradrenergic venlafaxine effect on the epididymal cauda region, focusing on aromatase, V-ATPase and EGF epithelial immunoexpression, smooth muscle (SM) integrity and mast cells number (MCN). Male rats were distributed into control (CG; n = 10) and venlafaxine (VFG, n = 10) groups. VFG received 30 mg/kg b.w. of venlafaxine for 35 days. The epididymal cauda was processed for light and transmission electron microscopy (TEM). The expression of connexin 43 (Cx43) and estrogen alpha (Esr1), adrenergic (Adra1a) and serotonergic (Htr1b) receptors were analyzed. Clear cells (CCs) area, SM thickness, viable spermatozoa (VS) and MCN were evaluated. Apoptosis was confirmed by TUNEL and TEM. The following immunoreactions were performed: T, aromatase, T/aromatase co-localization, V-ATPase, EGF, Cx43 and PCNA. The increased Adra1a and reduced Htr1b expressions confirmed the noradrenergic and serotonergic venlafaxine effects, respectively, corroborating the increased MCN, apoptosis and atrophy of SM. In VFG, the epithelial EGF increased, explaining Cx43 overexpression and basal cells mitotic activity. T aromatization and Esr1 downregulation indicate high estrogen levels, explaining CCs hypertrophy and changes in the V-ATPase localization, corroborating VS reduction. Thus, in addition to serotonergic/noradrenergic effects, T/estrogen imbalance, induced by venlafaxine, impairs epididymal structure and function.
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Epidídimo , ATPasas de Translocación de Protón Vacuolares , Ratas , Masculino , Animales , Clorhidrato de Venlafaxina/farmacología , Clorhidrato de Venlafaxina/metabolismo , Aromatasa , Conexina 43/metabolismo , Mastocitos/metabolismo , Factor de Crecimiento Epidérmico/metabolismo , ATPasas de Translocación de Protón Vacuolares/metabolismo , ATPasas de Translocación de Protón Vacuolares/farmacología , Estrógenos/farmacología , Miocitos del Músculo Liso/metabolismoRESUMEN
Submandibular gland (SMG) is responsive to androgens via androgen receptor (AR). We verified whether cimetidine induces androgenic dysfunction in SMG, and evaluated the structural integrity, cell death and immunoexpression of actin, EGF and V-ATPase in androgen-deficient SMG. Male rats received cimetidine (CMTG) and control animals (CG) received saline. Granular convoluted tubules (GCTs) diameter and number of acinar cell nuclei were evaluated. TUNEL and immunofluorescence reactions for detection of AR, testosterone, actin, EGF and V-ATPase were quantitatively analysed. In CG, testosterone immunolabelling was detected in acinar and ductal cells cytoplasm. AR-immunolabelled nuclei were observed in acinar cells whereas ductal cells showed AR-immunostained cytoplasm, indicating a non-genomic AR action. In CMTG, the weak testosterone and AR immunoexpression confirmed cimetidine-induced androgenic failure. A high cell death index was correlated with decreased number of acinar cells, GCTs diameter and EGF immunoexpression under androgenic dysfunction. Actin immunofluorescence decreased in the SMG cells, but an increased and diffuse cytoplasmic V-ATPase immunolabelling was observed in striated ducts, suggesting a disruption in the actin-dependent V-ATPase recycling due to androgenic failure. Our findings reinforce the androgenic role in the maintenance of SMG histophysiology, and point to a potential clinical use of cimetidine against androgen-dependent glandular tumour cells.
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Cimetidina/uso terapéutico , Inhibidores del Citocromo P-450 CYP1A2/uso terapéutico , Receptores Androgénicos/metabolismo , Glándula Submandibular/efectos de los fármacos , Actinas/metabolismo , Animales , Cimetidina/farmacología , Inhibidores del Citocromo P-450 CYP1A2/farmacología , Evaluación Preclínica de Medicamentos , Factor de Crecimiento Epidérmico/metabolismo , Masculino , Ratas Sprague-Dawley , Glándula Submandibular/metabolismo , Testosterona/metabolismo , ATPasas de Translocación de Protón Vacuolares/metabolismoRESUMEN
KEY MESSAGE: Transgenic sugarcane expressing V-ATPase subunit E dsRNA affects growth and survival of Sphenophorus levis. Plants being sessile organisms are constantly confronted with several biotic and abiotic stresses. Sugarcane (Saccharum spp) is a major tropical crop widely cultivated for its sugar and other by-products. In Brazil, sugarcane plantations account for significant production losses due to Sphenophorus levis (sugarcane weevil) infestations. With the existing control measures being less effective, there arises a necessity for advanced strategies. Our bioassay injection experiments with V-ATPase E dsRNA in S. levis larvae showed significant mortality and reduction in transcription levels. Furthermore, we down-regulated the V-ATPase E gene of S. levis in transgenic sugarcane using an RNAi approach. The resultant RNAi transgenic lines exhibited reduction in larval growth and survival, without compromising plant performance under controlled environment. Our results illustrate that RNAi-mediated down-regulation of key genes is a promising approach in imparting resistance to sugarcane weevil.
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Saccharum/genética , ATPasas de Translocación de Protón Vacuolares/genética , Gorgojos/crecimiento & desarrollo , Animales , Animales Modificados Genéticamente , Quimera , Expresión Génica , Control de Insectos , Proteínas de Insectos/genética , Larva , Plantas Modificadas Genéticamente , Interferencia de ARN , ARN Bicatenario/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Saccharum/fisiología , Gorgojos/genéticaRESUMEN
BACKGROUND: Granular cell tumors (GCTs) are rare neuroectodermal soft tissue neoplasms that mainly affect the skin of the upper limbs and trunks and the oral cavity. GCTs are derived from Schwann cells and, ultrastructurally, their intracytoplasmic granules are considered autophagosomes or autophagolysosomes and are consistent with myelin accumulation. METHODS: In this study, a convenience set of 22 formalin-fixed, paraffin-embedded samples of oral GCTs, all but one sample located at the tongue, was screened for mutations by whole-exome (WES) or targeted sequencing. RESULTS: WES revealed two novel variants in genes of the vacuolar ATPase (V-ATPase) complex: ATP6AP1 frameshift c.746_749del, leading to p.P249Hfs*4, and ATP6V1A non-synonymous c.G868A, leading to p.D290N. Each of these mutations occurred in one case. With regard to the samples that were wild type for these V-ATPase variants, at least two samples presented variants in genes that are part of endosomal/lysosomal/autophagosomal networks including ABCA8, ABCC6, AGAP3, ATG9A, CTSB, DNAJC13, GALC, NPC1, SLC15A3, SLC31A2, and TMEM104. CONCLUSION: Although the mechanisms involved in oral GCT initiation and progression remain unclear, our results suggest that oral GCTs have V-ATPase variants similarly to GCTs from other tissues/organs, and additionally show variants in lysosomes/endosomes/autophagosomal genes.
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Tumor de Células Granulares , ATPasas de Translocación de Protón Vacuolares , Biología , Tumor de Células Granulares/genética , Humanos , Lisosomas , ATPasas de Translocación de Protón Vacuolares/genética , Secuenciación del ExomaRESUMEN
Over the past few years, the use of RNA interference (RNAi) for insect pest management has attracted considerable interest in academia and industry as a pest-specific and environment-friendly strategy for pest control. For the success of this technique, the presence of core RNAi genes and a functional silencing machinery is essential. Therefore, the aim of this study was to test whether the Neotropical brown stinkbug Euschistus heros has the main RNAi core genes and whether the supply of dsRNA could generate an efficient gene silencing response. To do this, total mRNA of all developmental stages was sequenced on an Illumina platform, followed by a de novo assembly, gene annotation and RNAi-related gene identification. Once RNAi-related genes were identified, nuclease activities in hemolymph were investigated through an ex vivo assay. To test the functionality of the siRNA machinery, E. heros adults were microinjected with ~28 ng per mg of insect of a dsRNA targeting the V-ATPase-A gene. Mortality, relative transcript levels of V-ATPase-A, and the expression of the genes involved in the siRNA machinery, Dicer-2 (DCR-2) and Argonaute 2 (AGO-2), were analyzed. Transcriptome sequencing generated more than 126 million sequenced reads, and these were annotated in approximately 80,000 contigs. The search of RNAi-related genes resulted in 47 genes involved in the three major RNAi pathways, with the absence of sid-like homologous. Although ex vivo incubation of dsRNA in E. heros hemolymph showed rapid degradation, there was 35% mortality at 4 days after treatment and a significant reduction in V-ATPase-A gene expression. These results indicated that although sid-like genes are lacking, the dsRNA uptake mechanism was very efficient. Also, 2-fold and 4-fold overexpression of DCR-2 and AGO-2, respectively, after dsRNA supply indicated the activation of the siRNA machinery. Consequently, E. heros has proven to be sensitive to RNAi upon injection of dsRNA into its hemocoel. We believe that this finding together with a publically available transcriptome and the validation of a responsive RNAi machinery provide a starting point for future field applications against one of the most important soybean pests in South America.
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Perfilación de la Expresión Génica/veterinaria , Hemípteros/crecimiento & desarrollo , ARN Interferente Pequeño/genética , ATPasas de Translocación de Protón Vacuolares/genética , Animales , Regulación del Desarrollo de la Expresión Génica , Hemípteros/genética , Secuenciación de Nucleótidos de Alto Rendimiento/veterinaria , Control de Insectos , Proteínas de Insectos/genética , Anotación de Secuencia Molecular , Análisis de Secuencia de ARN/veterinaria , América del SurRESUMEN
BACKGROUND: V-ATPases are hetero-oligomeric enzymes consisting of 13 subunits and playing key roles in ion homeostasis and signaling. Differential expression of these proton pumps has been implicated in carcinogenesis and metastasis. To elucidate putative molecular signatures underlying these phenomena, we evaluated the expression of V-ATPase genes in esophageal squamous cell carcinoma (ESCC) and extended the analysis to other cancers. METHODS: Expression of all V-ATPase genes were analyzed in ESCC by a microarray data and in different types of tumors available from public databases. Expression of C isoforms was validated by qRT-PCR in paired ESCC samples. FINDINGS: A differential expression pattern of V-ATPase genes was found in different tumors, with combinations in up- and down-regulation leading to an imbalance in the expression ratios of their isoforms. Particularly, a high C1 and low C2 expression pattern accurately discriminated ESCC from normal tissues. Structural modeling of C2a isoform uncovered motifs for oncogenic kinases in an additional peptide stretch, and an actin-biding domain downstream to this sequence. INTERPRETATION: Altogether these data revealed that the expression ratios of subunits/isoforms could form a conformational code that controls the H+ pump regulation and interactions related to tumorigenesis. This study establishes a paradigm change by uncovering multi-cancer molecular signatures present in the V-ATPase structure, from which future studies must address the complexity of the onco-related V-ATPase assemblies as a whole, rather than targeting changes in specific subunit isoforms. FUNDING: This work was supported by grants from CNPq and FAPERJ-Brazil.
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Neoplasias Esofágicas/enzimología , Neoplasias Esofágicas/genética , Subunidades de Proteína/metabolismo , ATPasas de Translocación de Protón Vacuolares/genética , Anciano , Secuencia de Aminoácidos , Biomarcadores de Tumor/metabolismo , Neoplasias Esofágicas/diagnóstico , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/metabolismo , Masculino , Persona de Mediana Edad , Modelos Moleculares , Subunidades de Proteína/química , Subunidades de Proteína/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Curva ROC , Homología Estructural de Proteína , ATPasas de Translocación de Protón Vacuolares/química , ATPasas de Translocación de Protón Vacuolares/metabolismoRESUMEN
The goal of this study was to investigate the immunolocalization of inositol 1,4,5-trisphosphate receptor (IP3R) and vacuolar ATPase (V-ATPase) in ameloblastomas with special attention to the invasive front. Thirty-seven cases of previously diagnosed formalin-fixed paraffin-embedded (FFPE) human ameloblastoma samples were selected for this study. The samples were grouped according to the predominant histologic pattern and comprised twelve plexiform, eighteen follicular, and seven unicystic ameloblastomas. Of the unicystic variants, six demonstrated purely luminal and intraluminal growth, and one displayed mural extension. One granular cell variant was included in the follicular ameloblastoma group. All specimens were evaluated for IP3R and V-ATPase expression by immunohistochemistry (IHC). IP3R was positive in columnar cells, similar to ameloblasts, and non-peripheral cells in all samples. In the area of tumor protrusion and front of invasion, membranous and cystoplasmic IP3R expression was observed. In contrast, areas adjacent to tumoral protrusion demonstrated only membranous staining patterns. V-ATPase was not expressed in peripheral columnar cells of the unicystic and granular cell variants of ameloblastoma; however, strong staining was present in these cells in plexiform ameloblastomas, follicular ameloblastomas, and areas of mural growth of unicystic ameloblastomas. In areas of tumor protrusion, reactivity for V-ATPase was observed with both membranous and cytoplasmic staining, while other areas showed only membranous V-ATPase. These findings suggest that concomitant immunolocalization of IP3R and V-ATPase, with both cytoplasmic and membranous expression in the peripheral columnar cells, may indicate the invasive potential of ameloblastomas. Furthermore, these results suggest the tumoral spread of ameloblastomas may be correlated with the autophagy process and channelopathy. The expression of these proteins could establish a baseline for future research and provide therapeutic targets for treatment of ameloblastomas.
Asunto(s)
Ameloblastoma/patología , Biomarcadores de Tumor/análisis , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Neoplasias Maxilomandibulares/patología , ATPasas de Translocación de Protón Vacuolares/metabolismo , Humanos , Inmunohistoquímica , Receptores de Inositol 1,4,5-Trifosfato/análisis , ATPasas de Translocación de Protón Vacuolares/análisisRESUMEN
Manganese is considered essential for animal growth. Manganese ions serve as cofactors to three mitochondrial enzymes: superoxide dismutase (Sod2), arginase and glutamine synthase, and to glycosyltransferases residing in the Golgi. In Drosophila melanogaster, manganese has also been implicated in the formation of ceramide phosphoethanolamine, the insect's sphingomyelin analogue, a structural component of cellular membranes. Manganese overload leads to neurodegeneration and toxicity in both humans and Drosophila. Here, we report specific absorption and accumulation of manganese during the first week of adulthood in flies, which correlates with an increase in Sod2 activity during the same period. To test the requirement of dietary manganese for this accumulation, we generated a Drosophila model of manganese deficiency. Due to the lack of manganese-specific chelators, we used chemically defined media to grow the flies and deplete them of the metal. Dietary manganese depletion reduced Sod2 activity. We then examined gene and protein expression changes in the intestines of manganese depleted flies. We found adaptive responses to the presumed loss of known manganese-dependent enzymatic activities: less glutamine synthase activity (amination of glutamate to glutamine) was compensated by 50% reduction in glutaminase (deamination of glutamine to glutamate); less glycosyltransferase activity, predicted to reduce protein glycosylation, was compensated by 30% reduction in lysosomal mannosidases (protein deglycosylating enzymes); less ceramide phosphoethanolamine synthase activity was compensated by 30% reduction in the Drosophila sphingomyeline phospodiesterase, which could catabolize ceramide phosphoethanolamine in flies. Reduced Sod2 activity, predicted to cause superoxide-dependent iron-sulphur cluster damage, resulted in cellular iron misregulation.
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Drosophila melanogaster/fisiología , Intestinos/fisiología , Manganeso/deficiencia , Animales , Dieta , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Iones/metabolismo , Manganeso/análisis , RNA-Seq , Superóxido Dismutasa/metabolismo , ATPasas de Translocación de Protón Vacuolares/metabolismoRESUMEN
X-linked myopathy with excessive autophagy (XMEA) is a genetic disease associated with weakness of the proximal muscles. It is caused by mutations in the VMA21 gene, coding for a chaperone that functions in the vacuolar ATPase (v-ATPase) assembly. Mutations associated with lower content of assembled v-ATPases lead to an increase in lysosomal pH, culminating in partial blockage of macroautophagy, with accumulation of vacuoles of undigested content. Here, we studied a 5-year-old boy affected by XMEA, caused by a small indel in the VMA21 gene. Detection of sarcoplasmic Lc3 (also known as MAP1LC3B)-positive vacuoles in his muscle biopsy confirmed an autophagy defect. To understand how autophagy is regulated in XMEA myogenesis, we used patient-derived muscle cells to evaluate autophagy during in vitro muscle differentiation. An increase in lysosomal pH was observed in the patient's cells, compatible with predicted functional defect of his mutation. Additionally, there was an increase in autophagic flux in XMEA myotubes. Interestingly, we observed that differentiation of XMEA myoblasts was altered, with increased myotube formation observed through a higher fusion index, which was not dependent on lysosomal acidification. Moreover, no variation in the expression of myogenic factors nor the presence of regenerating fibers in the patient's muscle were observed. Myoblast fusion is a tightly regulated process; therefore, the uncontrolled fusion of XMEA myoblasts might generate cells that are not as functional as normal muscle cells. Our data provide new evidence on the reason for predominant muscle involvement in the context of the XMEA phenotype.This article has an associated First Person interview with the first author of the paper.
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Diferenciación Celular , Enfermedades Genéticas Ligadas al Cromosoma X/patología , Músculo Esquelético/patología , Enfermedades Musculares/patología , Autofagia , Secuencia de Bases , Biopsia , Brasil , Proliferación Celular , Preescolar , Femenino , Regulación de la Expresión Génica , Enfermedades Genéticas Ligadas al Cromosoma X/genética , Humanos , Recién Nacido , Lisosomas/metabolismo , Masculino , Fusión de Membrana , Desarrollo de Músculos/genética , Fibras Musculares Esqueléticas/metabolismo , Fibras Musculares Esqueléticas/patología , Enfermedades Musculares/genética , Mioblastos/metabolismo , Mioblastos/patología , Linaje , ARN Mensajero/genética , ARN Mensajero/metabolismo , ATPasas de Translocación de Protón Vacuolares/genética , ATPasas de Translocación de Protón Vacuolares/metabolismo , Vacuolas/patología , Vacuolas/ultraestructuraRESUMEN
INTRODUCTION: Malignant Infantile Osteopetrosis (MIOP) is a rare and severe genetic disorder due to abnormal osteoclast activity. OBJECTIVE: To report an infant who presented Malignant Infantile Osteopetrosis, reviewing the most relevant diagnostic and therapeutic aspects. CLINICAL CASE: A ten- month-old male infant with diagnosis of MIOP confirmed after presenting thrombocytopenia and visceromegaly. He was the first child of non-consanguineous parents, and among the findings, he presented severe hepatosplenomegaly, thrombocytopenia, and anemia; visual and hearing impair ment, and repeated infections. The diagnosis was confirmed by genetic study, which identified two heterozygous mutations in the TCIRG1 gene. Hematopoietic stem cells were transplanted without hematological recovery. The patient died due to occlusive venous disease. DISCUSSION: MIOP is a rare, severe, and early-onset disease, with a high rate of suspicion necessary in the presence of hepa- tosplenomegaly and bone marrow failure. Early diagnosis and hematopoietic stem cells transplanta tion are the only potentially therapeutic interventions of this lethal entity.
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Trasplante de Células Madre Hematopoyéticas/métodos , Osteopetrosis/diagnóstico , ATPasas de Translocación de Protón Vacuolares/genética , Resultado Fatal , Humanos , Lactante , Masculino , Mutación , Osteopetrosis/genética , Osteopetrosis/fisiopatologíaRESUMEN
INTRODUCTION: Idiopathic pulmonary fibrosis (IPF) is a lethal disease with an unclear pathogenic mechanism. Components of the renin-angiotensin system (RAS) have a role in the pathogenesis of IPF, specifically, the aspartyl protease renin acts as a profibrotic factor in the lung. However, the concentration of the RAS components renin and soluble (pro)renin receptor (sPRR) have not been previously evaluated neither in serum nor in bronchoalveolar lavage fluid (BAL) of patients with IPF or chronic Hypersensitivity pneumonitis (cHP), a disease which may be confused with IPF. METHODS: The serum levels of renin [IPF patients (n = 70), cHP patients (n = 83), and controls (n = 26)] and sPRR [IPF (n = 28), cHP (37), and controls (n = 20)] were measured by ELISA. Renin was also quantified in BALs of IPF patients and controls by Western blot. RESULTS: We found that the levels of renin were higher in serum samples from IPF patients when compared with cHP patients and controls. Furthermore, BALs from IPF patients had more renin than BALs from controls. Unlike renin, the serum levels of sPRR were lower in IPF and cHP patients than in control individuals. CONCLUSIONS: The high levels of renin in sera and BALs of IPF patients suggest that renin might play a major role in the pathogenesis of IPF. Results from BAL confirm that renin is produced locally in the lung. Serum levels of renin could be used to differentiate IPF from cHP.
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Alveolitis Alérgica Extrínseca/metabolismo , Fibrosis Pulmonar Idiopática/metabolismo , Receptores de Superficie Celular/sangre , Renina/metabolismo , ATPasas de Translocación de Protón Vacuolares/sangre , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Alveolitis Alérgica Extrínseca/fisiopatología , Western Blotting , Líquido del Lavado Bronquioalveolar/química , Enfermedad Crónica , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Fibrosis Pulmonar Idiopática/fisiopatología , Masculino , Persona de Mediana Edad , Oxígeno/metabolismo , Capacidad de Difusión Pulmonar , Capacidad Vital , Adulto JovenRESUMEN
INTRODUCCIÓN: Osteopetrosis Infantil Maligna (OIM) es un grave e inusual desorden genético debi do a una actividad osteoclástica anormal. OBJETIVO: Reportar lactante en quien se documentó una Osteopetrosis Infantil Maligna, revisando aspectos diagnósticos y terapéuticos más relevantes. CASO CLÍNICO: Reportamos un lactante de 10 meses de sexo masculino en quien se confirmó OIM tras presentar plaquetopenia y visceromegalias. En su historial destacó ser primer hijo de padres no consanguíneos, y entre sus hallazgos presentó hepatoesplenomegalia, plaquetopenia y anemia graves, compromiso sensorial visual y auditivo e infecciones a repetición. El diagnóstico fue confirmado mediante estudio genético, el cual identificó 2 mutaciones heterocigotas en el gen TCIRG1. Se rea lizó trasplante de precursores hematopoyéticos, sin haber presentado recuperación hematológica, falleciendo por enfermedad veno oclusiva. DISCUSIÓN: La OIM es una enfermedad inusual, grave y de inicio temprano, siendo necesario un elevado índice de sospecha ante hepatoesplenomegalia y falla medular. El diagnóstico temprano y el trasplante de precursores hematopoyéticos son las únicas intervenciones potencialmente curativas de esta entidad letal.
INTRODUCCIÓN: Malignant Infantile Osteopetrosis (MIOP) is a rare and severe genetic disorder due to abnormal osteoclast activity. OBJECTIVE: To report an infant who presented Malignant Infantile Osteopetrosis, reviewing the most relevant diagnostic and therapeutic aspects. CLINICAL CASE: A ten- month-old male infant with diagnosis of MIOP confirmed after presenting thrombocytopenia and visceromegaly. He was the first child of non-consanguineous parents, and among the findings, he presented severe hepatosplenomegaly, thrombocytopenia, and anemia; visual and hearing impairment, and repeated infections. The diagnosis was confirmed by genetic study, which identified two heterozygous mutations in the TCIRG1 gene. Hematopoietic stem cells were transplanted without hematological recovery. The patient died due to occlusive venous disease. DISCUSSION: MIOP is a rare, severe, and early-onset disease, with a high rate of suspicion necessary in the presence of hepatosplenomegaly and bone marrow failure. Early diagnosis and hematopoietic stem cells transplanta tion are the only potentially therapeutic interventions of this lethal entity.
Asunto(s)
Humanos , Masculino , Lactante , Osteopetrosis/diagnóstico , Trasplante de Células Madre Hematopoyéticas/métodos , ATPasas de Translocación de Protón Vacuolares/genética , Osteoporosis/fisiopatología , Osteoporosis/genética , Resultado Fatal , MutaciónRESUMEN
V-ATPases are part of the membrane components of pathogen-containing vacuoles, although their function in intracellular infection remains elusive. In addition to organelle acidification, V-ATPases are alternatively implicated in membrane fusion and anti-inflammatory functions controlled by ATP6V0d2, the d subunit variant of the V-ATPase complex. Therefore, we evaluated the role of ATP6V0d2 in the biogenesis of pathogen-containing vacuoles using ATP6V0d2 knock-down macrophages infected with the protozoan parasite Leishmania amazonensis. These parasites survive within IFNγ/LPS-activated inflammatory macrophages, multiplying in large/fusogenic parasitophorous vacuoles (PVs) and inducing ATP6V0d2 upregulation. ATP6V0d2 knock-down decreased macrophage cholesterol levels and inhibited PV enlargement without interfering with parasite multiplication. However, parasites required ATP6V0d2 to resist the influx of oxidized low-density lipoprotein (ox-LDL)-derived cholesterol, which restored PV enlargement in ATP6V0d2 knock-down macrophages by replenishing macrophage cholesterol pools. Thus, we reveal parasite-mediated subversion of host V-ATPase function toward cholesterol retention, which is required for establishing an inflammation-resistant intracellular parasite niche.
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Colesterol/metabolismo , Regulación Enzimológica de la Expresión Génica , Homeostasis , Leishmania/metabolismo , Macrófagos/metabolismo , Regulación hacia Arriba , ATPasas de Translocación de Protón Vacuolares/biosíntesis , Vacuolas/metabolismo , Animales , Lipoproteínas LDL/metabolismo , Macrófagos/parasitología , Macrófagos/patología , Ratones , Ratones Endogámicos BALB C , Células RAW 264.7 , Vacuolas/parasitología , Vacuolas/patologíaRESUMEN
BACKGROUND: Distal renal tubular acidosis (dRTA) is a kidney tubulopathy that causes a state of normal anion gap metabolic acidosis due to impairment of urine acidification. This review aims to summarize the etiology, pathophysiology, clinical findings, diagnosis and therapeutic approach of dRTA, with emphasis on genetic causes of dRTA. DATA SOURCES: Literature reviews and original research articles from databases, including PubMed and Google Scholar. Manual searching was performed to identify additional studies about dRTA. RESULTS: dRTA is characterized as the dysfunction of the distal urinary acidification, leading to metabolic acidosis. In pediatric patients, the most frequent etiology of dRTA is the genetic alteration of genes responsible for the codification of distal tubule channels, whereas, in adult patients, dRTA is more commonly secondary to autoimmune diseases, use of medications and uropathies. Patients with dRTA exhibit failure to thrive and important laboratory alterations, which are used to define the diagnosis. The oral alkali and potassium supplementation can correct the biochemical defects, improve clinical manifestations and avoid nephrolithiasis and nephrocalcinosis. CONCLUSIONS: dRTA is a multifactorial disease leading to several clinical manifestations. Clinical and laboratory alterations can be corrected by alkali replacement therapy.
Asunto(s)
Acidosis Tubular Renal/tratamiento farmacológico , Acidosis Tubular Renal/genética , Acidosis Tubular Renal/diagnóstico , Acidosis Tubular Renal/fisiopatología , Adolescente , Proteína 1 de Intercambio de Anión de Eritrocito/genética , Niño , Humanos , Mutación , ATPasas de Translocación de Protón Vacuolares/genéticaRESUMEN
AIMS: Pregnancy is a physiological stage with profound cardiovascular changes leading to hypotension. Preeclampsia (PE) reverts these normal changes inducing hypertension. Renin-angiotensin system (RAS) has been related in PE genesis. It has been reported a novel receptor in the system, the Prorenin/Renin receptor (PRR), with several roles in renal and cardiovascular illnesses. It is not known, however, if PRR changes its expression or is activated during normal or PE-complicated pregnancy on tissues intimately related to hypertension. So, the aim of this work was to describe PRR expression during normal and hypertensive pregnancy in rats. METHODS: We used a subrenal aortic coarctation (SRAC) model in rats. Atria, septum and ventricular heart tissue, aorta and renal tissue samples were homogenized and immunoblotted using anti-PRR and anti-PLZF antibodies. We also measured gene expression by RT-PCR. KEY FINDINGS: Hypertension and proteinuria were observed in SRAC-pregnant rats. In pregnant, non-SRAC rats, PRR showed a higher expression of both, gene and protein compared to non-pregnant rats in heart, aorta and kidney tissues. PE induces a very high expression of PRR in cardiac tissues and, on the contrary, decreases PRR expression in both, aorta and kidney. PLZF, a marker of PRR function, was augmented only in aorta and kidney in non-SRAC pregnant rats. In SRAC-pregnant rats, PLZF increment disappeared. SIGNIFICANCE: These findings indicate that PRR expression changes differently during pregnancy and PE in tissues related to cardiovascular functions and suggest a probable participation of the receptor during normal and preeclamptic pregnancy in the rat.
Asunto(s)
Hipertensión Inducida en el Embarazo/fisiopatología , Preeclampsia/fisiopatología , Proteinuria/fisiopatología , Receptores de Superficie Celular/genética , Sistema Renina-Angiotensina , Animales , Aorta/metabolismo , Modelos Animales de Enfermedad , Femenino , Regulación de la Expresión Génica , Hipertensión Inducida en el Embarazo/genética , Riñón/metabolismo , Preeclampsia/genética , Embarazo , Ratas , Ratas Wistar , Receptores de Superficie Celular/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , ATPasas de Translocación de Protón VacuolaresRESUMEN
BACKGROUND: Metastatic tumor cells have acidic extracellular pH and differential electrochemical H+ gradients generated across their cell membranes by V-type H+-ATPases. This study shows that inhibition of the V-ATPases by the plant-derived monoterpene Myrtenal results in tumor cell death and decreased metastatic dissemination in mice. METHODS: The Myrtenal anticancer toxicity was evaluated in vitro using murine (B16F0 and B16F10) and human (SkMel-5) melanoma cell lines, and in in vivo mouse metastatic dissemination model. Proton flux and extracellular acidification were directly evaluated at the surface of living cells using a non-invasive selective ion electrode approach. RESULTS: The inhibition of V-ATPases by 100⯵M Myrtenal disrupted the electrochemical H+ gradient across the cell membranes, strongly induced cell death (4-5 fold), and decreased tumor cells migration and invasion in vitro. Myrtenal (15â¯mg/kg) also significantly reduced metastasis induced by B16F10 in vivo, further reinforcing that V-ATPase is a molecular target to halt the progression of cancers. CONCLUSIONS: These data revealed the therapeutic potential of Myrtenal as inhibitor of melanoma progression proposing a mechanism of action by which once inhibited by this monoterpene the proton pumps fail to activate cancer-related differential electrochemical gradients and H+ fluxes across the tumor cell membranes, disrupting pH signatures inherent in tumor progression, resulting in reprogrammed cell death and metastasis inhibition. GENERAL SIGNIFICANCE: The work represents a new mechanistic strategy for contention of melanoma, the most aggressive and deadly form of cutaneous neoplasm, and highlights Myrtenal, other related monoterpenes and derivatives as promising proton pump inhibitors with high chemotherapeutic potential.
Asunto(s)
Antineoplásicos/farmacología , Melanoma/patología , Neoplasias Cutáneas/patología , Terpenos/farmacología , ATPasas de Translocación de Protón Vacuolares/antagonistas & inhibidores , Animales , Monoterpenos Bicíclicos , Muerte Celular , Línea Celular Tumoral , Membrana Celular/metabolismo , Movimiento Celular , Modelos Animales de Enfermedad , Resistencia a Antineoplásicos , Electrodos , Humanos , Concentración de Iones de Hidrógeno , Masculino , Melanoma/tratamiento farmacológico , Melanoma Experimental , Ratones , Ratones Endogámicos C57BL , Invasividad Neoplásica , Metástasis de la Neoplasia/tratamiento farmacológico , Metástasis de la Neoplasia/prevención & control , Protones , Neoplasias Cutáneas/tratamiento farmacológico , ATPasas de Translocación de Protón Vacuolares/metabolismoRESUMEN
Background: Autosomal malignant osteopetrosis is a rare condition arising from dysfunction of bone-resorbing osteoclasts, in which diagnosis requires a high suspicion index. Treatment of choice is allogeneic stem cell transplantation. Best outcomes occur if the procedure is carried out before damage to cranial nerves ensues; nonetheless, patients improve their clinical condition. Case report: An 8-month-old infant was referred for hematology consultation for cytopenias, hepatomegaly, and growth failure. Autosomal malignant osteopetrosis was diagnosed on the basis of physical findings, alteration in calcium and phosphorus metabolism, and hyperdensity of bone. DNA was obtained from the patient and parents; compound heterozygosity of the TCIRG1 gene with a previously non-described deletion (c.1809_1818del) was identified. Conclusions: A new pathogenic mutation of TCIRG1 was identified in a Mexican osteopetrotic patient. Hematopoietic stem cell transplantation was offered as the best available treatment but declined by the parents. An early recognition and wider access to this procedure should be implemented.
Introducción: La osteopetrosis infantil maligna es una condición rara cuyo origen es la deficiente reabsorción ósea por parte de los osteoclastos. Su diagnóstico requiere un alto índice de sospecha. El tratamiento de elección es el trasplante alogénico de células hematopoyéticas. Los mejores desenlaces ocurren si el procedimiento se lleva a cabo antes de que ocurra daño a los nervios craneales. Caso clínico: Paciente masculino de 8 meses de edad fue referido a la consulta de hematología por citopenias, hepatomegalia y falla para crecer. Se diagnosticó osteopetrosis infantil maligna basándose en los hallazgos de la exploración física, la alteración del metabolismo del calcio y el fósforo y la hiperdensidad del hueso. Se obtuvo ADN del paciente y ambos padres; se demostró un heterocigosidad compuesta del gen TCIRG1 con una deleción (c.1809_1818del) no descrita previamente. Conclusiones: Una nueva mutación patogénica de TCIRG1 se identificó en un paciente mexicano con osteopetrosis. Se ofreció trasplante de células progenitoras hematopoyéticas como el mejor tratamiento disponible, pero fue rechazado por los padres. Se necesita un reconocimiento temprano y la implementación del acceso generalizado a este procedimiento.
Asunto(s)
Trasplante de Células Madre Hematopoyéticas , Osteopetrosis/congénito , ATPasas de Translocación de Protón Vacuolares/genética , Humanos , Lactante , Masculino , México , Mutación , Osteopetrosis/diagnóstico , Osteopetrosis/genética , Osteopetrosis/terapia , Eliminación de Secuencia , Negativa del Paciente al TratamientoRESUMEN
Abstract Background: Autosomal malignant osteopetrosis is a rare condition arising from dysfunction of bone-resorbing osteoclasts, in which diagnosis requires a high suspicion index. Treatment of choice is allogeneic stem cell transplantation. Best outcomes occur if the procedure is carried out before damage to cranial nerves ensues; nonetheless, patients improve their clinical condition. Case report: An 8-month-old infant was referred for hematology consultation for cytopenias, hepatomegaly, and growth failure. Autosomal malignant osteopetrosis was diagnosed on the basis of physical findings, alteration in calcium and phosphorus metabolism, and hyperdensity of bone. DNA was obtained from the patient and parents; compound heterozygosity of the TCIRG1 gene with a previously non-described deletion (c.1809_1818del) was identified. Conclusions: A new pathogenic mutation of TCIRG1 was identified in a Mexican osteopetrotic patient. Hematopoietic stem cell transplantation was offered as the best available treatment but declined by the parents. An early recognition and wider access to this procedure should be implemented.
Resumen Introducción: La osteopetrosis infantil maligna es una condición rara cuyo origen es la deficiente reabsorción ósea por parte de los osteoclastos. Su diagnóstico requiere un alto índice de sospecha. El tratamiento de elección es el trasplante alogénico de células hematopoyéticas. Los mejores desenlaces ocurren si el procedimiento se lleva a cabo antes de que ocurra daño a los nervios craneales. Caso clínico: Paciente masculino de 8 meses de edad fue referido a la consulta de hematología por citopenias, hepatomegalia y falla para crecer. Se diagnosticó osteopetrosis infantil maligna basándose en los hallazgos de la exploración física, la alteración del metabolismo del calcio y el fósforo y la hiperdensidad del hueso. Se obtuvo ADN del paciente y ambos padres; se demostró un heterocigosidad compuesta del gen TCIRG1 con una deleción (c.1809_1818del) no descrita previamente. Conclusiones: Una nueva mutación patogénica de TCIRG1 se identificó en un paciente mexicano con osteopetrosis. Se ofreció trasplante de células progenitoras hematopoyéticas como el mejor tratamiento disponible, pero fue rechazado por los padres. Se necesita un reconocimiento temprano y la implementación del acceso generalizado a este procedimiento.