RESUMEN
The Japanese archipelago is a terminal location for human migration, and the contemporary Japanese people represent a unique population whose genomic diversity has been shaped by multiple migrations from Eurasia. We analyzed the genomic characteristics that define the genetic makeup of the modern Japanese population from a population genetics perspective from the genomic data of 9,287 samples obtained by high-coverage whole-genome sequencing (WGS) by the National Center Biobank Network. The dataset comprised populations from the Ryukyu Islands and other parts of the Japanese archipelago (Hondo). The Hondo population underwent two episodes of population decline during the Jomon period, corresponding to the Late Neolithic, and the Edo period, corresponding to the Early Modern era, while the Ryukyu population experienced a population decline during the shell midden period of the Late Neolithic in this region. Haplotype analysis suggested increased allele frequencies for genes related to alcohol and fatty acid metabolism, which were reported as loci that had experienced positive natural selection. Two genes related to alcohol metabolism were found to be 12,500 years out of phase with the time when they began to increase in the allele frequency; this finding indicates that the genomic diversity of Japanese people has been shaped by events closely related to agriculture and food production.
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Pueblos del Este de Asia , Genética de Población , Humanos , Variación Genética , Japón , Secuenciación Completa del Genoma , Pueblos del Este de Asia/genéticaRESUMEN
Androgen(s) is one of the sex steroids that are involved in many physiological phenomena of vertebrate species. Although androgens were originally identified as male sex hormones, it is well known now that they are also essential in females. As in the case of other steroid hormones, androgen is produced from cholesterol through serial enzymatic reactions. Although testis is a major tissue to produce androgens in all species, androgens are also produced in ovary and adrenal (interrenal tissue). Testosterone is the most common and famous androgen. It represents a major androgen both in males and females of almost vertebrate species. In addition, testosterone is a precursor for producing significant androgens such as11-ketotestosterone, 5α-dihydrotestosterone, 11-ketodihydrotestosterones and 15α-hydroxytestosterone in a species- or sex-dependent manner for their homeostasis. In this article, we will review the significance and characteristics of these androgens, following a description of the history of testosterone discovery and its synthetic pathways.
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Andrógenos , Testosterona , Masculino , Animales , Femenino , Ovario , Testículo , VertebradosRESUMEN
Repeated implantation failure is a major cause of infertility among healthy women. Uterine ß-catenin (CTNNB1) plays a critical role in implantation. However, the role of embryonic CTNNB1 during implantation remains unclear. We addressed this topic by analyzing mice carrying Ctnnb1-deficient (Ctnnb1Δ/Δ) embryos. Ctnnb1Δ/Δ embryos were produced by intercrossing mice bearing Ctnnb1-deficient eggs and sperms. We found that Ctnnb1Δ/Δ embryos developed to the blastocyst stage; thereafter, they were resorbed, leaving empty decidual capsules. Moreover, leukemia inhibitory factor, a uterine factor essential for implantation, was undetectable in Ctnnb1Δ/Δ blastocysts. Furthermore, CDX2, a transcription factor that determines the fate of trophectoderm cells, was not observed in Ctnnb1Δ/Δ blastocysts. Intrauterine injection with uterine fluids (from control mice) and recombinant mouse leukemia inhibitory factor proteins rescued the uterine response to Ctnnb1Δ/Δ blastocysts. These results suggest that embryonic CTNNB1 is required for the secretion of blastocyst-derived factor(s) that open the implantation window, indicating that the uterine response to implantation can be induced using supplemental materials. Therefore, our results may contribute to the discovery of a similar mechanism in humans, leading to a better understanding of the pathogenesis of repeated implantation failure.
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Implantación del Embrión , beta Catenina , Animales , Femenino , Humanos , Ratones , beta Catenina/genética , beta Catenina/metabolismo , Blastocisto/metabolismo , Implantación del Embrión/fisiología , Factor Inhibidor de Leucemia/genética , Factor Inhibidor de Leucemia/metabolismo , Útero/metabolismoRESUMEN
For patients who are often embarrassed and uncomfortable when exposing their breasts and having them touched by physicians of different genders during auscultation, we are developing a robotic system that performs auscultation over clothing. As the technical issue, the sound obtained through the clothing is often attenuated. This study aims to investigate clothing-induced acoustic attenuation and develop a suppression method for it. Because the attenuation is due to the loss of energy as sound propagates through a medium with viscosity, we hypothesized that the attenuation is improved by compressing clothing and shortening the sound propagation distance. Then, the amplitude spectrum of the heart sound was obtained over clothes of different thicknesses and materials in a phantom study and human trial at varying contact forces with a developed passive-actuated end-effector. Our results demonstrate the feasibility of the attenuation suppression method by applying an optimum contact force, which varied according to the clothing condition. In the phantom experiments, the attenuation rate was improved maximumly by 48% when applying the optimal contact force (1 N). In human trials, the attenuation rate was under the acceptable attenuation (40%) when applying the optimal contact force in all combinations in each subject. The proposed method promises the potential of robotic auscultation toward eliminating gender bias.
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Procedimientos Quirúrgicos Robotizados , Masculino , Humanos , Femenino , Sexismo , Acústica , Auscultación , VestuarioRESUMEN
JARID2 is an accessory component of Polycomb repressive complex-2 (PRC2) required for the differentiation of embryonic stem cells (ESCs). A role for JARID2 in the recruitment of PRC2 to target genes silenced during differentiation has been put forward, but the molecular details remain unclear. We identified a 30-amino-acid region of JARID2 that mediates interactions with long noncoding RNAs (lncRNAs) and found that the presence of lncRNAs stimulated JARID2-EZH2 interactions in vitro and JARID2-mediated recruitment of PRC2 to chromatin in vivo. Native and crosslinked RNA immunoprecipitations of JARID2 revealed that Meg3 and other lncRNAs from the imprinted Dlk1-Dio3 locus, an important regulator of development, interacted with PRC2 via JARID2. Lack of MEG3 expression in human induced pluripotent cells altered the chromatin distribution of JARID2, PRC2, and H3K27me3. Our findings show that lncRNAs facilitate JARID2-PRC2 interactions on chromatin and suggest a mechanism by which lncRNAs contribute to PRC2 recruitment.
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Cromatina/metabolismo , Complejo Represivo Polycomb 2/metabolismo , Complejo Represivo Polycomb 2/fisiología , ARN no Traducido/metabolismo , Animales , Sitios de Unión , Células Cultivadas , Proteína Potenciadora del Homólogo Zeste 2 , Células HEK293 , Humanos , Células Madre Pluripotentes Inducidas , Ratones , Complejo Represivo Polycomb 2/química , ARN Largo no Codificante/metabolismoRESUMEN
Dilated cardiomyopathy (DCM) is caused by various gene variants and characterized by systolic dysfunction. Lamin variants have been reported to have a poor prognosis. Medical and device therapies are not sufficient to improve the prognosis of DCM with the lamin variants. Recently, induced pluripotent stem (iPS) cells have been used for research on genetic disorders. However, few studies have evaluated the contractile function of cardiac tissue with lamin variants. The aim of this study was to elucidate the function of cardiac cell sheet tissue derived from patients with lamin variant DCM. iPS cells were generated from a patient with lamin A/C (LMNA) -mutant DCM (LMNA p.R225X mutation). After cardiac differentiation and purification, cardiac cell sheets that were fabricated through cultivation on a temperature-responsive culture dish were transferred to the surface of the fibrin gel, and the contractile force was measured. The contractile force and maximum contraction velocity, but not the maximum relaxation velocity, were significantly decreased in cardiac cell sheet tissue with the lamin variant. A qRT-PCR analysis revealed that mRNA expression of some contractile proteins, cardiac transcription factors, Ca2+-handling genes, and ion channels were downregulated in cardiac tissue with the lamin variant.Human iPS-derived bioengineered cardiac tissue with the LMNA p.R225X mutation has the functional properties of systolic dysfunction and may be a promising tissue model for understanding the underlying mechanisms of DCM.
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Cardiomiopatías , Cardiomiopatía Dilatada , Células Madre Pluripotentes Inducidas , Cardiomiopatías/metabolismo , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Mutación , Miocitos Cardíacos/metabolismoRESUMEN
Ornithine transcarbamylase deficiency (OTCD) is a metabolic and genetic disease caused by dysfunction of the hepatocytic urea cycle. To develop new drugs or therapies for OTCD, it is ideal to use models that are more closely related to human metabolism and pathology. Primary human hepatocytes (HHs) isolated from two patients (a 6-month-old boy and a 5-year-old girl) and a healthy donor were transplanted into host mice (hemi-, hetero-OTCD mice, and control mice, respectively). HHs were isolated from these mice and used for serial transplantation into the next host mouse or for in vitro experiments. Histological, biochemical, and enzyme activity analyses were performed. Cultured HHs were treated with ammonium chloride or therapeutic drugs. Replacement rates exceeded 80% after serial transplantation in both OTCD mice. These highly humanized OTCD mice showed characteristics similar to OTCD patients that included increased blood ammonia levels and urine orotic acid levels enhanced by allopurinol. Hemi-OTCD mice showed defects in OTC expression and significantly low enzymatic activities, while hetero-OTCD mice showed residual OTC expression and activities. A reduction in ammonium metabolism was observed in cultured HHs from OTCD mice, and treatment with the therapeutic drug reduced the ammonia levels in the culture medium. In conclusion, we established in vivo OTC mouse models with hemi- and hetero-patient HHs. HHs isolated from the mice were useful as an in vitro model of OTCD. These OTC models could be a source of valuable patient-derived hepatocytes that would enable large scale and reproducible experiments using the same donor.
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Hepatocitos/trasplante , Enfermedad por Deficiencia de Ornitina Carbamoiltransferasa/terapia , Ornitina Carbamoiltransferasa/genética , Amoníaco/sangre , Animales , Preescolar , Modelos Animales de Enfermedad , Femenino , Regulación de la Expresión Génica , Hepatocitos/química , Hepatocitos/citología , Humanos , Lactante , Masculino , Ratones , Enfermedad por Deficiencia de Ornitina Carbamoiltransferasa/genética , Ácido Orótico/orinaRESUMEN
Genetically engineered pigs play an indispensable role in the study of rare monogenic diseases. Pigs harboring a gene responsible for a specific disease can be efficiently generated via somatic cell cloning. The generation of somatic cell-cloned pigs from male cells with mutation(s) in an X chromosomal gene is a reliable and straightforward method for reproducing X-linked genetic diseases (XLGDs) in pigs. However, the severe symptoms of XLGDs are often accompanied by impaired growth and reproductive disorders, which hinder the reproduction of these valuable model animals. Here, we generated unique chimeric boars composed of mutant cells harboring a lethal XLGD and normal cells. The chimeric boars exhibited the cured phenotype with fertility while carrying and transmitting the genotype of the XLGD. This unique reproduction system permits routine production of XLGD model pigs through the male-based breeding, thereby opening an avenue for translational research using disease model pigs.
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Técnicas de Cultivo de Embriones/métodos , Enfermedades Genéticas Ligadas al Cromosoma X/genética , Reproducción/genética , Animales , Animales Modificados Genéticamente/genética , Cruzamiento , Quimera , Clonación de Organismos/métodos , Modelos Animales de Enfermedad , Fertilidad , Técnicas de Inactivación de Genes/métodos , Ingeniería Genética/métodos , Masculino , Técnicas de Transferencia Nuclear , Porcinos/genéticaRESUMEN
The hedgehog signaling pathway is a vital factor for embryonic development and stem cell maintenance. Dysregulation of its function results in tumor initiation and progression. The aim of this research was to establish a disease model of hedgehog-related tumorigenesis with Gorlin syndrome-derived induced pluripotent stem cells (GS-iPSCs). Induced neural progenitor cells from GS-iPSCs (GS-NPCs) show constitutive high GLI1 expression and higher sensitivity to smoothened (SMO) inhibition compared with wild-type induced neural progenitor cells (WT-NPCs). The differentiation process from iPSCs to NPCs may have similarity in gene expression to Hedgehog signal-related carcinogenesis. Therefore, GS-NPCs may be useful for screening compounds to find effective drugs to control Hedgehog signaling activity.
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Síndrome del Nevo Basocelular , Células-Madre Neurales , Transducción de Señal/genética , Receptor Smoothened , Proteína con Dedos de Zinc GLI1 , Anilidas , Síndrome del Nevo Basocelular/genética , Síndrome del Nevo Basocelular/metabolismo , Diferenciación Celular/genética , Células Cultivadas , Humanos , Modelos Biológicos , Células-Madre Neurales/citología , Células-Madre Neurales/metabolismo , Receptor Patched-1 , Piridinas , Receptor Smoothened/genética , Receptor Smoothened/metabolismo , Proteína con Dedos de Zinc GLI1/antagonistas & inhibidores , Proteína con Dedos de Zinc GLI1/genética , Proteína con Dedos de Zinc GLI1/metabolismoRESUMEN
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
RESUMEN
Men and women become infertile with age, but the mechanism of declining male fertility, more specifically, the decrease in in sperm quality, is not well known. Citrate synthase (CS) is a core enzyme of the mitochondrial tricarboxylic acid (TCA) cycle, which directly controls cellular function. Extra-mitochondrial CS (eCS) is produced and abundant in the sperm head; however, its role in male fertility is unknown. We investigated the role of eCS in male fertility by producing eCs-deficient (eCs-KO) mice. The initiation of the first spike of Ca2+ oscillation was substantially delayed in egg fused with eCs-KO sperm, despite normal expression of sperm factor phospholipase C zeta 1. The eCs-KO male mice were initially fertile, but the fertility dropped with age. Metabolomic analysis of aged sperm revealed that the loss of eCS enhances TCA cycle in the mitochondria with age, presumably leading to depletion of extra-mitochondrial citrate. The data suggest that eCS suppresses age-dependent male infertility, providing insights into the decline of male fertility with age.
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Envejecimiento/metabolismo , Señalización del Calcio/fisiología , Citrato (si)-Sintasa , Infertilidad Masculina/metabolismo , Espermatozoides , Animales , Citrato (si)-Sintasa/genética , Citrato (si)-Sintasa/metabolismo , Ciclo del Ácido Cítrico/fisiología , Femenino , Infertilidad Masculina/fisiopatología , Masculino , Metaboloma/fisiología , Ratones , Óvulo/metabolismo , Espermatozoides/enzimología , Espermatozoides/metabolismoRESUMEN
The objective was to prepare guidelines to perform the current optimum treatment by organizing effective and efficient treatments of hemangiomas and vascular malformations, confirming the safety, and systematizing treatment, employing evidence-based medicine (EBM) techniques and aimed at improvement of the outcomes. Clinical questions (CQs) were decided based on the important clinical issues. For document retrieval, key words for literature searches were set for each CQ and literature published from 1980 to the end of September 2014 was searched in Pubmed, Cochrane Library, and Japana Centra Revuo Medicina (JCRM). The strengths of evidence and recommendations acquired by systematic reviews were determined following the Medical Information Network Distribution System (MINDS) technique. A total of 33 CQs were used to compile recommendations and the subjects included efficacy of resection, sclerotherapy/embolization, drug therapy, laser therapy, radiotherapy, and other conservative treatment, differences in appropriate treatment due to the location of lesions and among symptoms, appropriate timing of treatment and tests, and pathological diagnosis deciding the diagnosis. Thus, the Japanese Clinical Practice Guidelines for Vascular Anomalies 2017 have been prepared as the evidence-based guidelines for the management of vascular anomalies.
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Hemangioma/terapia , Malformaciones Vasculares/terapia , Malformaciones Arteriovenosas/terapia , Embolización Terapéutica/métodos , Medicina Basada en la Evidencia , Humanos , Terapia por Láser/métodos , Escleroterapia/métodos , Resultado del TratamientoRESUMEN
Inner and middle ear disorders are the leading cause of hearing loss, and are said to be among the greatest risk factors of dementia. The use of regenerative medicine for the treatment of inner ear disorders may offer a potential alternative to cochlear implants for hearing recovery. In this paper, we reviewed recent research and clinical applications in middle and inner ear regeneration and cell therapy. Recently, the mechanism of inner ear regeneration has gradually been elucidated. "Inner ear stem cells," which may be considered the precursors of various cells in the inner ear, have been discovered in the cochlea and vestibule. Research indicates that cells such as hair cells, neurons, and spiral ligaments may form promising targets for inner ear regenerative therapies by the transplantation of stem cells, including mesenchymal stem cells. In addition, it is necessary to develop tests for the clinical monitoring of cell transplantation. Real-time imaging techniques and hearing rehabilitation techniques are also being investigated, and cell therapy has found clinical application in cochlear implant techniques.
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Oído Interno/fisiopatología , Pérdida Auditiva/fisiopatología , Pérdida Auditiva/terapia , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas/citología , Regeneración , Animales , Pérdida Auditiva/complicaciones , Humanos , Degeneración Nerviosa/complicacionesRESUMEN
Hermaphroditic invertebrates and plants have a self-recognition system on the cell surface of sperm and eggs, which prevents their self-fusion and enhances non-self-fusion, thereby contributing to genetic variation. However, the system of sperm-egg recognition in mammals is under debate. To address this issue, we explored the role of major histocompatibility complex class I (MHC class I, also known as histocompatibility 2-Kb or H2-Kb and H2-Db in mice) antigens by analyzing H2-Kb-/-H2-Db-/-ß2-microglobulin (ß2M)-/- triple-knockout (T-KO) male mice with full fertility. T-KO sperm exhibited an increased sperm number in the perivitelline space of wild-type (WT) eggs in vitro. Moreover, T-KO sperm showed multiple fusion with zona pellucida (ZP)-free WT eggs, implying that the ability of polyspermy block for sperm from T-KO males was weakened in WT eggs. When T-KO male mice were intercrossed with WT female mice, the percentage of females in progeny increased. We speculate that WT eggs prefer fusion with T-KO sperm, more specifically X-chromosome-bearing sperm (X sperm), suggesting the presence of preferential (non-random) fertilization in mammals, including humans.
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Fertilidad/genética , Antígenos de Histocompatibilidad Clase I/genética , Óvulo/metabolismo , Razón de Masculinidad , Interacciones Espermatozoide-Óvulo/genética , Espermatozoides/metabolismo , Animales , Proteínas de Unión al ADN/deficiencia , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/inmunología , Femenino , Fertilización In Vitro , Regulación de la Expresión Génica , Antígenos de Histocompatibilidad Clase I/inmunología , Humanos , Masculino , Ratones , Ratones Noqueados , Óvulo/citología , Recuento de Espermatozoides , Espermatozoides/citología , Microglobulina beta-2/deficiencia , Microglobulina beta-2/genética , Microglobulina beta-2/inmunologíaRESUMEN
Down syndrome is the most frequent chromosomal abnormality among live-born infants. All Down syndrome patients have mental retardation and are prone to develop early onset Alzheimer's disease. However, it has not yet been elucidated whether there is a correlation between the phenotype of Down syndrome and the extra chromosome 21. In this study, we continuously cultivated induced pluripotent stem cells (iPSCs) with chromosome 21 trisomy for more than 70 weeks, and serendipitously obtained revertant cells with normal chromosome 21 diploids from the trisomic cells during long-term cultivation. Repeated experiments revealed that this trisomy rescue was not due to mosaicism of chromosome 21 diploid cells and occurred at an extremely high frequency. We herewith report the spontaneous correction from chromosome 21 trisomy to disomy without genetic manipulation, chemical treatment or exposure to irradiation. The revertant diploid cells will possibly serve a reference for drug screening and a raw material of regenerative medicinal products for cell-based therapy.
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Síndrome de Down/genética , Células Madre Pluripotentes Inducidas , Células Cultivadas , Femenino , Humanos , Embarazo , TrisomíaRESUMEN
Tetraspanin CD9 is essential for sperm-egg fusion and also contributes to uterine repair through microexosome formation. Microexosomes share CD9 with exosomes and are released from eggs and uterine epithelial cells. However, the mechanism for the formation of microexosomes remains unknown. To address this issue, we examined membrane localization and extracellular release of CD9 proteins using uterine epithelial cells and secretions in mice and humans. In mice, CD9 localized predominantly on the basal region of the plasma membrane and relocated to the apical region upon embryo implantation. Furthermore, extracellular CD9 proteins were detected in uterine secretions of mice and women undergoing infertility treatment, but were below detectable levels in supernatants of pluripotent stem cells. Ultrastructural analysis demonstrated that membrane projections were shortened and the number of mitochondria was reduced in uterine epithelial cells lacking Cd9 genes. Our results suggest that CD9 repositioning and release affect both membrane structures and mitochondrial state in the uterus, and contribute to female fertility.
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Tetraspanina 29 , Útero , Animales , Secreciones Corporales/química , Secreciones Corporales/citología , Línea Celular , Ciclo Estral , Exosomas/química , Exosomas/metabolismo , Femenino , Humanos , Infertilidad Femenina , Ratones , Ratones Endogámicos C57BL , Mitocondrias/química , Mitocondrias/metabolismo , Tetraspanina 29/química , Tetraspanina 29/metabolismo , Tetraspanina 29/fisiología , Útero/química , Útero/citología , Útero/metabolismo , Útero/fisiologíaRESUMEN
SHOX resides in the short arm pseudoautosomal region (PAR1) of the sex chromosomes and escapes X inactivation. SHOX haploinsufficiency underlies idiopathic short stature (ISS) and Leri-Weill dyschondrosteosis (LWD). A substantial percentage of cases with SHOX haploinsufficiency arise from pseudoautosomal copy number variations (CNVs) involving putative enhancer regions of SHOX. Our previous study using peripheral blood samples showed that some CpG dinucleotides adjacent to SHOX exon 1 were hypomethylated in a healthy woman and methylated in a woman with gross X chromosomal rearrangements. However, it remains unknown whether submicroscopic pseudoautosomal CNVs cause aberrant DNA methylation of SHOX-flanking CpG islands. In this study, we examined the DNA methylation status of SHOX-flanking CpG islands in 50 healthy individuals and 10 ISS/LWD patients with pseudoautosomal CNVs. In silico analysis detected 3 CpG islands within the 20-kb region from the translation start site of SHOX. Pyrosequencing and bisulfite sequencing of genomic DNA samples revealed that these CpG islands were barely methylated in peripheral blood cells and cultured chondrocytes of healthy individuals, as well as in peripheral blood cells of ISS/LWD patients with pseudoautosomal CNVs. These results, in conjunction with our previous findings, indicate that the DNA methylation status of SHOX-flanking CpG islands can be affected by gross X-chromosomal abnormalities, but not by submicroscopic CNVs in PAR1. Such CNVs likely disturb SHOX expression through DNA methylation-independent mechanisms, which need to be determined in future studies.
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Metilación de ADN , Enfermedades Genéticas Ligadas al Cromosoma X/genética , Trastornos del Crecimiento/genética , Osteocondrodisplasias/genética , Proteína de la Caja Homeótica de Baja Estatura/genética , Adolescente , Adulto , Estudios de Casos y Controles , Células Cultivadas , Niño , Preescolar , Condrocitos , Islas de CpG , Variaciones en el Número de Copia de ADN , Femenino , Humanos , Análisis de Secuencia de ADNRESUMEN
In bacteria, a polymer of inorganic phosphate (Pi) (inorganic polyphosphate; polyP) is enzymatically produced and consumed as an alternative phosphate donor for adenosine triphosphate (ATP) production to protect against nutrient starvation. In vertebrates, polyP has been dismissed as a "molecular fossil" due to the lack of any known physiological function. Here, we have explored its possible role by producing transgenic (TG) mice widely expressing Saccharomyces cerevisiae exopolyphosphatase 1 (ScPPX1), which catalyzes hydrolytic polyP degradation. TG mice were produced and displayed reduced mitochondrial respiration in muscles. In female TG mice, the blood concentration of lactic acid was enhanced, whereas ATP storage in liver and brain tissues was reduced significantly. Thus, we suggested that the elongation of polyP reduces the intracellular Pi concentration, suppresses anaerobic lactic acid production, and sustains mitochondrial respiration. Our results provide an insight into the physiological role of polyP in mammals, particularly in females.
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Ácido Anhídrido Hidrolasas/metabolismo , Ácido Láctico/metabolismo , Fosfatos/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Respiración de la Célula/fisiología , Escherichia coli/metabolismo , Fermentación , Ácido Láctico/análisis , Ácido Láctico/sangre , Ratones , Ratones Transgénicos , Mitocondrias/metabolismo , Oocitos/metabolismo , Polímeros , Polifosfatos/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismoRESUMEN
Cyclooxygenase 2 (COX-2) is an inducible rate-limiting enzyme for prostanoid production. Because COX-2 represents one of the inducible genes in mouse mesenchymal stem cells upon differentiation into Leydig cells, we investigated COX-2 expression and production of prostaglandin (PG) in Leydig cells. Although COX-2 was undetectable in mouse testis, it was transiently induced in Leydig cells by human chorionic gonadotropin (hCG) administration. Consistent with the finding that Leydig cells expressed aldo-keto reductase 1B7 (PGF synthase) and PGE synthase 2, induction of COX-2 by hCG caused a marked increase in testicular PGF 2α and PGE 2 levels. Using mouse Leydig cell tumor-derived MA-10 cells as a model, it was indicated by reporter assays and electron mobility shift assays that transcription of the COX-2 gene was activated by CCAAT/enhancer-binding protein ß (C/EBPß) with cAMP-stimulation. C/EBPß expression was induced by cAMP-stimulation, whereas expression of C/EBP homolog protein (CHOP) was robustly downregulated. Transfection of CHOP expression plasmid inhibited cAMP-induced COX-2 promoter activity. In addition, CHOP reduced constitutive COX-2 expression in other mouse Leydig cell tumor-derived TM3 cells. These results indicate that COX-2 is induced in Leydig cells by activation of C/EBPß via reduction of CHOP expression upon gonadotropin-stimulation to produce PGF 2α and PGE 2 .
Asunto(s)
Proteína beta Potenciadora de Unión a CCAAT/metabolismo , Gonadotropina Coriónica/farmacología , Ciclooxigenasa 2/metabolismo , Dinoprostona/metabolismo , Células Intersticiales del Testículo/metabolismo , Sustancias para el Control de la Reproducción/farmacología , Animales , Línea Celular Tumoral , AMP Cíclico/metabolismo , Ciclooxigenasa 2/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Regiones Promotoras Genéticas , Transducción de Señal/efectos de los fármacos , Factor de Transcripción CHOP/genética , Factor de Transcripción CHOP/metabolismo , Transcripción Genética , TransfecciónRESUMEN
In female mammals, activation of Xist (X-inactive specific transcript) is essential for establishment of X chromosome inactivation. During early embryonic development in mice, paternal Xist is preferentially expressed whereas maternal Xist (Xm-Xist) is silenced. Unlike autosomal imprinted genes, Xist imprinting for Xm-Xist silencing was erased in cloned or parthenogenetic but not fertilized embryos. However, the molecular mechanism underlying the variable nature of Xm-Xist imprinting is poorly understood. Here, we revealed that Xm-Xist silencing depends on chromatin condensation states at the Xist/Tsix genomic region and on Rnf12 expression levels. In early preimplantation, chromatin decondensation via H3K9me3 loss and histone acetylation gain caused Xm-Xist derepression irrespective of embryo type. Although the presence of the paternal genome during pronuclear formation impeded Xm-Xist derepression, Xm-Xist was robustly derepressed when the maternal genome was decondensed before fertilization. Once Xm-Xist was derepressed by chromatin alterations, the derepression was stably maintained and rescued XmXpΔ lethality, indicating that loss of Xm-Xist imprinting was irreversible. In late preimplantation, Oct4 served as a chromatin opener to create transcriptional permissive states at Xm-Xist/Tsix genomic loci. In parthenogenetic embryos, Rnf12 overdose caused Xm-Xist derepression via Xm-Tsix repression; physiological Rnf12 levels were essential for Xm-Xist silencing maintenance in fertilized embryos. Thus, chromatin condensation and fine-tuning of Rnf12 dosage were crucial for Xist imprint maintenance by silencing Xm-Xist.