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1.
Cell ; 186(26): 5910-5924.e17, 2023 12 21.
Artículo en Inglés | MEDLINE | ID: mdl-38070509

RESUMEN

The evolution and development of the head have long captivated researchers due to the crucial role of the head as the gateway for sensory stimuli and the intricate structural complexity of the head. Although significant progress has been made in understanding head development in various vertebrate species, our knowledge of early human head ontogeny remains limited. Here, we used advanced whole-mount immunostaining and 3D imaging techniques to generate a comprehensive 3D cellular atlas of human head embryogenesis. We present detailed developmental series of diverse head tissues and cell types, including muscles, vasculature, cartilage, peripheral nerves, and exocrine glands. These datasets, accessible through a dedicated web interface, provide insights into human embryogenesis. We offer perspectives on the branching morphogenesis of human exocrine glands and unknown features of the development of neurovascular and skeletomuscular structures. These insights into human embryology have important implications for understanding craniofacial defects and neurological disorders and advancing diagnostic and therapeutic strategies.


Asunto(s)
Embrión de Mamíferos , Cabeza , Humanos , Morfogénesis , Cabeza/crecimiento & desarrollo
2.
Physiol Rev ; 100(3): 1349-1414, 2020 07 01.
Artículo en Inglés | MEDLINE | ID: mdl-32031468

RESUMEN

The male genital tract (MGT) is the target of a number of viral infections that can have deleterious consequences at the individual, offspring, and population levels. These consequences include infertility, cancers of male organs, transmission to the embryo/fetal development abnormalities, and sexual dissemination of major viral pathogens such as human immunodeficiency virus (HIV) and hepatitis B virus. Lately, two emerging viruses, Zika and Ebola, have additionally revealed that the human MGT can constitute a reservoir for viruses cleared from peripheral circulation by the immune system, leading to their sexual transmission by cured men. This represents a concern for future epidemics and further underlines the need for a better understanding of the interplay between viruses and the MGT. We review here how viruses, from ancient viruses that integrated the germline during evolution through old viruses (e.g., papillomaviruses originating from Neanderthals) and more modern sexually transmitted infections (e.g., simian zoonotic HIV) to emerging viruses (e.g., Ebola and Zika) take advantage of genital tract colonization for horizontal dissemination, viral persistence, vertical transmission, and endogenization. The MGT immune responses to viruses and the impact of these infections are discussed. We summarize the latest data regarding the sources of viruses in semen and the complex role of this body fluid in sexual transmission. Finally, we introduce key animal findings that are relevant for our understanding of viral infection and persistence in the human MGT and suggest future research directions.


Asunto(s)
Enfermedades Transmisibles Emergentes/virología , Genitales Masculinos/virología , Virosis/virología , Humanos , Masculino , Virosis/patología
3.
Diabetologia ; 67(6): 1066-1078, 2024 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-38630142

RESUMEN

AIMS/HYPOTHESIS: Rodent pancreas development has been described in great detail. On the other hand, there are still gaps in our understanding of the developmental trajectories of pancreatic cells during human ontogenesis. Here, our aim was to map the spatial and chronological dynamics of human pancreatic cell differentiation and proliferation by using 3D imaging of cleared human embryonic and fetal pancreases. METHODS: We combined tissue clearing with light-sheet fluorescence imaging in human embryonic and fetal pancreases during the first trimester of pregnancy. In addition, we validated an explant culture system enabling in vitro proliferation of pancreatic progenitors to determine the mitogenic effect of candidate molecules. RESULTS: We detected the first insulin-positive cells as early as five post-conceptional weeks, two weeks earlier than previously observed. We observed few insulin-positive clusters at five post-conceptional weeks (mean ± SD 9.25±5.65) with a sharp increase to 11 post-conceptional weeks (4307±152.34). We identified a central niche as the location of onset of the earliest insulin cell production and detected extra-pancreatic loci within the adjacent developing gut. Conversely, proliferating pancreatic progenitors were located in the periphery of the epithelium, suggesting the existence of two separated pancreatic niches for differentiation and proliferation. Additionally, we observed that the proliferation ratio of progenitors ranged between 20% and 30%, while for insulin-positive cells it was 1%. We next unveiled a mitogenic effect of the platelet-derived growth factor AA isoform (PDGFAA) in progenitors acting through the pancreatic mesenchyme by increasing threefold the number of proliferating progenitors. CONCLUSIONS/INTERPRETATION: This work presents a first 3D atlas of the human developing pancreas, charting both endocrine and proliferating cells across early development.


Asunto(s)
Diferenciación Celular , Proliferación Celular , Imagenología Tridimensional , Páncreas , Humanos , Páncreas/embriología , Páncreas/citología , Páncreas/metabolismo , Diferenciación Celular/fisiología , Femenino , Células Madre/citología , Células Madre/metabolismo , Embarazo , Insulina/metabolismo
4.
Reprod Biol Endocrinol ; 22(1): 60, 2024 May 22.
Artículo en Inglés | MEDLINE | ID: mdl-38778396

RESUMEN

BACKGROUND: Reproduction in women is at risk due to exposure to chemicals that can disrupt the endocrine system during different windows of sensitivity throughout life. Steroid hormone levels are fundamental for the normal development and function of the human reproductive system, including the ovary. This study aims to elucidate steroidogenesis at different life-stages in human ovaries. METHODS: We have developed a sensitive and specific LC-MS/MS method for 21 important steroid hormones and measured them at different life stages: in media from cultures of human fetal ovaries collected from elective terminations of normally progressing pregnancy and in media from adult ovaries from Caesarean section patients, and follicular fluid from women undergoing infertility treatment. Statistically significant differences in steroid hormone levels and their ratios were calculated with parametric tests. Principal component analysis (PCA) was applied to explore clustering of the ovarian-derived steroidogenic profiles. RESULTS: Comparison of the 21 steroid hormones revealed clear differences between the various ovarian-derived steroid profiles. Interestingly, we found biosynthesis of both canonical and "backdoor" pathway steroid hormones and corticosteroids in first and second trimester fetal and adult ovarian tissue cultures. 17α-estradiol, a less potent naturally occurring isomer of 17ß-estradiol, was detected only in follicular fluid. PCA of the ovarian-derived profiles revealed clusters from: adult ovarian tissue cultures with relatively high levels of androgens; first trimester and second trimester fetal ovarian tissue cultures with relatively low estrogen levels; follicular fluid with the lowest androgens, but highest corticosteroid, progestogen and estradiol levels. Furthermore, ratios of specific steroid hormones showed higher estradiol/ testosterone and estrone/androstenedione (indicating higher CYP19A1 activity, p < 0.01) and higher 17-hydroxyprogesterone/progesterone and dehydroepiandrosterone /androstenedione (indicating higher CYP17A1 activity, p < 0.01) in fetal compared to adult ovarian tissue cultures. CONCLUSIONS: Human ovaries demonstrate de novo synthesis of non-canonical and "backdoor" pathway steroid hormones and corticosteroids. Elucidating the steroid profiles in human ovaries improves our understanding of physiological, life-stage dependent, steroidogenic capacity of ovaries and will inform mechanistic studies to identify endocrine disrupting chemicals that affect female reproduction.


Asunto(s)
Feto , Ovario , Humanos , Femenino , Ovario/metabolismo , Adulto , Embarazo , Feto/metabolismo , Hormonas Esteroides Gonadales/biosíntesis , Hormonas Esteroides Gonadales/metabolismo , Hormonas Esteroides Gonadales/análisis , Espectrometría de Masas en Tándem , Líquido Folicular/metabolismo , Líquido Folicular/química , Estradiol/metabolismo , Cromatografía Liquida
5.
Arch Toxicol ; 97(3): 849-863, 2023 03.
Artículo en Inglés | MEDLINE | ID: mdl-36653537

RESUMEN

Exposure to endocrine-disrupting chemicals (EDCs) during development may cause reproductive disorders in women. Although female reproductive endpoints are assessed in rodent toxicity studies, a concern is that typical endpoints are not sensitive enough to detect chemicals of concern to human health. If so, measured endpoints must be improved or new biomarkers of effects included. Herein, we have characterized the dynamic transcriptional landscape of developing rat ovaries exposed to two well-known EDCs, diethylstilbestrol (DES) and ketoconazole (KTZ), by 3' RNA sequencing. Rats were orally exposed from day 7 of gestation until birth, and from postnatal day 1 until days 6, 14 or 22. Three exposure doses for each chemical were used: 3, 6 and 12 µg/kg bw/day of DES; 3, 6, 12 mg/kg bw/day of KTZ. The transcriptome changed dynamically during perinatal development in control ovaries, with 1137 differentially expressed genes (DEGs) partitioned into 3 broad expression patterns. A cross-species deconvolution strategy based on a mouse ovary developmental cell atlas was used to map any changes to ovarian cellularity across the perinatal period to allow for characterization of actual changes to gene transcript levels. A total of 184 DEGs were observed across dose groups and developmental stages in DES-exposed ovaries, and 111 DEGs in KTZ-exposed ovaries across dose groups and developmental stages. Based on our analyses, we have identified new candidate biomarkers for female reproductive toxicity induced by EDC, including Kcne2, Calb2 and Insl3.


Asunto(s)
Disruptores Endocrinos , Canales de Potasio con Entrada de Voltaje , Humanos , Embarazo , Ratones , Femenino , Ratas , Animales , Dietilestilbestrol/toxicidad , Ovario , Disruptores Endocrinos/toxicidad , Cetoconazol , Reproducción , Canales de Potasio con Entrada de Voltaje/farmacología
6.
FASEB J ; 35(7): e21718, 2021 07.
Artículo en Inglés | MEDLINE | ID: mdl-34105801

RESUMEN

Acetaminophen, aspirin, and ibuprofen are mild analgesics commonly used by pregnant women, the sole current recommendation being to avoid ibuprofen from the fifth month of gestation. The nephrotoxicity of these three analgesics is well documented in adults, as is their interference with prostaglandins biosynthesis. Here we investigated the effect of these analgesics on human first trimester kidneys ex vivo. We first evaluated prostaglandins biosynthesis functionality by performing a wide screening of prostaglandin expression patterns in first trimester human kidneys. We demonstrated that prostaglandins biosynthesis machinery is functional during early nephrogenesis. Human fetal kidney explants aged 7-12 developmental weeks were exposed ex vivo to ibuprofen, aspirin or acetaminophen for 7 days, and analyzed by histology, immunohistochemistry, and flow cytometry. This study has revealed that these analgesics induced a spectrum of abnormalities within early developing structures, ranging from cell death to a decline in differentiating glomeruli density. These results warrant caution for the use of these medicines during the first trimester of pregnancy.


Asunto(s)
Analgésicos/efectos adversos , Feto/efectos de los fármacos , Glomérulos Renales/efectos de los fármacos , Organogénesis/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Femenino , Feto/metabolismo , Humanos , Glomérulos Renales/metabolismo , Embarazo , Primer Trimestre del Embarazo/efectos de los fármacos , Prostaglandinas/metabolismo
7.
Proc Natl Acad Sci U S A ; 115(4): E715-E724, 2018 01 23.
Artículo en Inglés | MEDLINE | ID: mdl-29311296

RESUMEN

Concern has been raised over increased male reproductive disorders in the Western world, and the disruption of male endocrinology has been suggested to play a central role. Several studies have shown that mild analgesics exposure during fetal life is associated with antiandrogenic effects and congenital malformations, but the effects on the adult man remain largely unknown. Through a clinical trial with young men exposed to ibuprofen, we show that the analgesic resulted in the clinical condition named "compensated hypogonadism," a condition prevalent among elderly men and associated with reproductive and physical disorders. In the men, luteinizing hormone (LH) and ibuprofen plasma levels were positively correlated, and the testosterone/LH ratio decreased. Using adult testis explants exposed or not exposed to ibuprofen, we demonstrate that the endocrine capabilities from testicular Leydig and Sertoli cells, including testosterone production, were suppressed through transcriptional repression. This effect was also observed in a human steroidogenic cell line. Our data demonstrate that ibuprofen alters the endocrine system via selective transcriptional repression in the human testes, thereby inducing compensated hypogonadism.


Asunto(s)
Analgésicos no Narcóticos/efectos adversos , Hipogonadismo/inducido químicamente , Ibuprofeno/efectos adversos , Hormona Luteinizante/sangre , Testosterona/sangre , Adulto , Analgésicos no Narcóticos/sangre , Línea Celular , Expresión Génica/efectos de los fármacos , Humanos , Hipogonadismo/sangre , Ibuprofeno/sangre , Técnicas In Vitro , Células Intersticiales del Testículo/efectos de los fármacos , Células Intersticiales del Testículo/metabolismo , Masculino , Persona de Mediana Edad , Prostaglandinas/biosíntesis , Células de Sertoli/efectos de los fármacos
8.
Int J Mol Sci ; 22(13)2021 Jun 22.
Artículo en Inglés | MEDLINE | ID: mdl-34206462

RESUMEN

Human fetal gonads acquire endocrine steroidogenic capabilities early during their differentiation. Genetic studies show that this endocrine function plays a central role in the sexually dimorphic development of the external genitalia during fetal development. When this endocrine function is dysregulated, congenital malformations and pathologies are the result. In this review, we explain how the current knowledge of steroidogenesis in human fetal gonads has benefited from both the technological advances in steroid measurements and the assembly of detailed knowledge of steroidogenesis machinery and its expression in human fetal gonads. We summarise how the conversion of radiolabelled steroid precursors, antibody-based assays, mass spectrometry, ultrastructural studies, and the in situ labelling of proteins and mRNA have all provided complementary information. In this review, our discussion goes beyond the debate on recommendations concerning the best choice between the different available technologies, and their degrees of reproducibility and sensitivity. The available technologies and techniques can be used for different purposes and, as long as all quality controls are rigorously employed, the question is how to maximise the generation of robust, reproducible data on steroid hormones and their crucial roles in human fetal development and subsequent functions.


Asunto(s)
Feto/metabolismo , Hormonas Esteroides Gonadales/metabolismo , Gónadas/metabolismo , Investigación , Femenino , Humanos , Inmunoensayo , Masculino , Espectrometría de Masas , Ovario/metabolismo , Ovario/ultraestructura , Investigación/tendencias , Desarrollo Sexual/genética , Testículo/metabolismo , Testículo/ultraestructura
9.
Hum Reprod ; 35(5): 1099-1119, 2020 05 01.
Artículo en Inglés | MEDLINE | ID: mdl-32412604

RESUMEN

STUDY QUESTION: Which transcriptional program triggers sex differentiation in bipotential gonads and downstream cellular events governing fetal testis and ovary development in humans? SUMMARY ANSWER: The characterization of a dynamically regulated protein-coding and non-coding transcriptional landscape in developing human gonads of both sexes highlights a large number of potential key regulators that show an early sexually dimorphic expression pattern. WHAT IS KNOWN ALREADY: Gonadal sex differentiation is orchestrated by a sexually dimorphic gene expression program in XX and XY developing fetal gonads. A comprehensive characterization of its non-coding counterpart offers promising perspectives for deciphering the molecular events underpinning gonad development and for a complete understanding of the etiology of disorders of sex development in humans. STUDY DESIGN, SIZE, DURATION: To further investigate the protein-coding and non-coding transcriptional landscape during gonad differentiation, we used RNA-sequencing (RNA-seq) and characterized the RNA content of human fetal testis (N = 24) and ovaries (N = 24) from 6 to 17 postconceptional week (PCW), a key period in sex determination and gonad development. PARTICIPANTS/MATERIALS, SETTING, METHODS: First trimester fetuses (6-12 PCW) and second trimester fetuses (13-14 and 17 PCW) were obtained from legally induced normally progressing terminations of pregnancy. Total RNA was extracted from whole human fetal gonads and sequenced as paired-end 2 × 50 base reads. Resulting sequences were mapped to the human genome, allowing for the assembly and quantification of corresponding transcripts. MAIN RESULTS AND THE ROLE OF CHANCE: This RNA-seq analysis of human fetal testes and ovaries at seven key developmental stages led to the reconstruction of 22 080 transcripts differentially expressed during testicular and/or ovarian development. In addition to 8935 transcripts displaying sex-independent differential expression during gonad development, the comparison of testes and ovaries enabled the discrimination of 13 145 transcripts that show a sexually dimorphic expression profile. The latter include 1479 transcripts differentially expressed as early as 6 PCW, including 39 transcription factors, 40 long non-coding RNAs and 20 novel genes. Despite the use of stringent filtration criteria (expression cut-off of at least 1 fragment per kilobase of exon model per million reads mapped, fold change of at least 2 and false discovery rate adjusted P values of less than <1%), the possibility of assembly artifacts and of false-positive differentially expressed transcripts cannot be fully ruled out. LARGE-SCALE DATA: Raw data files (fastq) and a searchable table (.xlss) containing information on genomic features and expression data for all refined transcripts have been submitted to the NCBI GEO under accession number GSE116278. LIMITATIONS, REASONS FOR CAUTION: The intrinsic nature of this bulk analysis, i.e. the sequencing of transcripts from whole gonads, does not allow direct identification of the cellular origin(s) of the transcripts characterized. Potential cellular dilution effects (e.g. as a result of distinct proliferation rates in XX and XY gonads) may account for a few of the expression profiles identified as being sexually dimorphic. Finally, transcriptome alterations that would result from exposure to pre-abortive drugs cannot be completely excluded. Although we demonstrated the high quality of the sorted cell populations used for experimental validations using quantitative RT-PCR, it cannot be totally excluded that some germline expression may correspond to cell contamination by, for example, macrophages. WIDER IMPLICATIONS OF THE FINDINGS: For the first time, this study has led to the identification of 1000 protein-coding and non-coding candidate genes showing an early, sexually dimorphic, expression pattern that have not previously been associated with sex differentiation. Collectively, these results increase our understanding of gonad development in humans, and contribute significantly to the identification of new candidate genes involved in fetal gonad differentiation. The results also provide a unique resource that may improve our understanding of the fetal origin of testicular and ovarian dysgenesis syndromes, including cryptorchidism and testicular cancers. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the French National Institute of Health and Medical Research (Inserm), the University of Rennes 1, the French School of Public Health (EHESP), the Swiss National Science Foundation [SNF n° CRS115_171007 to B.J.], the French National Research Agency [ANR n° 16-CE14-0017-02 and n° 18-CE14-0038-02 to F.C.], the Medical Research Council [MR/L010011/1 to P.A.F.] and the European Community's Seventh Framework Programme (FP7/2007-2013) [under grant agreement no 212885 to P.A.F.] and from the European Union's Horizon 2020 Research and Innovation Programme [under grant agreement no 825100 to P.A.F. and S.M.G.]. There are no competing interests related to this study.


Asunto(s)
Diferenciación Sexual , Testículo , Femenino , Feto , Gónadas , Humanos , Masculino , Ovario , Embarazo , Diferenciación Sexual/genética
10.
Arch Toxicol ; 94(10): 3359-3379, 2020 10.
Artículo en Inglés | MEDLINE | ID: mdl-32638039

RESUMEN

Modern living challenges female reproductive health. We are witnessing a rise in reproductive disorders and drop in birth rates across the world. The reasons for these manifestations are multifaceted and most likely include continuous exposure to an ever-increasing number of chemicals. The cause-effect relationships between chemical exposure and female reproductive disorders, however, have proven problematic to determine. This has made it difficult to assess the risks chemical exposures pose to a woman's reproductive development and function. To address this challenge, this review uses the adverse outcome pathway (AOP) concept to summarize current knowledge about how chemical exposure can affect female reproductive health. We have a special focus on effects on the ovaries, since they are essential for lifelong reproductive health in women, being the source of both oocytes and several reproductive hormones, including sex steroids. The AOP framework is widely accepted as a new tool for toxicological safety assessment that enables better use of mechanistic knowledge for regulatory purposes. AOPs equip assessors and regulators with a pragmatic network of linear cause-effect relationships, enabling the use of a wider range of test method data in chemical risk assessment and regulation. Based on current knowledge, we propose ten putative AOPs relevant for female reproductive disorders that can be further elaborated and potentially be included in the AOPwiki. This effort is an important step towards better safeguarding the reproductive health of all girls and women.


Asunto(s)
Rutas de Resultados Adversos , Seguridad Química , Exposición Materna , Ovario/efectos de los fármacos , Salud Reproductiva , Animales , Enfermedades del Sistema Endocrino/inducido químicamente , Femenino , Humanos , Ratones , Enfermedades del Ovario/inducido químicamente , Ovario/fisiopatología , Embarazo , Medición de Riesgo , Pruebas de Toxicidad
11.
Int J Mol Sci ; 21(9)2020 May 01.
Artículo en Inglés | MEDLINE | ID: mdl-32370092

RESUMEN

Currently available test methods are not well-suited for the identification of chemicals that disturb hormonal processes involved in female reproductive development and function. This renders women's reproductive health at increasing risk globally, which, coupled with increasing incidence rates of reproductive disorders, is of great concern. A woman's reproductive health is largely established during embryonic and fetal development and subsequently matures during puberty. The endocrine system influences development, maturation, and function of the female reproductive system, thereby making appropriate hormone levels imperative for correct functioning of reproductive processes. It is concerning that the effects of human-made chemicals on the endocrine system and female reproductive health are poorly addressed in regulatory chemical safety assessment, partly because adequate test methods are lacking. Our EU-funded project FREIA aims to address this need by increasing understanding of how endocrine disrupting chemicals (EDCs) can impact female reproductive health. We will use this information to provide better test methods that enable fit-for-purpose chemical regulation and then share our knowledge, promote a sustainable society, and improve the reproductive health of women globally.


Asunto(s)
Disruptores Endocrinos/farmacología , Reproducción/efectos de los fármacos , Salud Reproductiva , Animales , Sistema Endocrino/efectos de los fármacos , Exposición a Riesgos Ambientales , Contaminantes Ambientales/efectos adversos , Femenino , Humanos , Pubertad/efectos de los fármacos , Medición de Riesgo , Factores de Riesgo
12.
Bioinformatics ; 34(12): 2116-2122, 2018 06 15.
Artículo en Inglés | MEDLINE | ID: mdl-29385404

RESUMEN

Motivation: At the same time that toxicologists express increasing concern about reproducibility in this field, the development of dedicated databases has already smoothed the path toward improving the storage and exchange of raw toxicogenomic data. Nevertheless, none provides access to analyzed and interpreted data as originally reported in scientific publications. Given the increasing demand for access to this information, we developed TOXsIgN, a repository for TOXicogenomic sIgNatures. Results: The TOXsIgN repository provides a flexible environment that facilitates online submission, storage and retrieval of toxicogenomic signatures by the scientific community. It currently hosts 754 projects that describe more than 450 distinct chemicals and their 8491 associated signatures. It also provides users with a working environment containing a powerful search engine as well as bioinformatics/biostatistics modules that enable signature comparisons or enrichment analyses. Availability and implementation: The TOXsIgN repository is freely accessible at http://toxsign.genouest.org. Website implemented in Python, JavaScript and MongoDB, with all major browsers supported. Supplementary information: Supplementary data are available at Bioinformatics online.


Asunto(s)
Bases de Datos Factuales , Programas Informáticos , Toxicogenética/métodos , Animales , Humanos
13.
PLoS Genet ; 10(5): e1004340, 2014 May.
Artículo en Inglés | MEDLINE | ID: mdl-24784881

RESUMEN

The Hedgehog (Hh) family of secreted proteins act as morphogens to control embryonic patterning and development in a variety of organ systems. Post-translational covalent attachment of cholesterol and palmitate to Hh proteins are critical for multimerization and long range signaling potency. However, the biological impact of lipid modifications on Hh ligand distribution and signal reception in humans remains unclear. In the present study, we report a unique case of autosomal recessive syndromic 46,XY Disorder of Sex Development (DSD) with testicular dysgenesis and chondrodysplasia resulting from a homozygous G287V missense mutation in the hedgehog acyl-transferase (HHAT) gene. This mutation occurred in the conserved membrane bound O-acyltransferase (MBOAT) domain and experimentally disrupted the ability of HHAT to palmitoylate Hh proteins such as DHH and SHH. Consistent with the patient phenotype, HHAT was found to be expressed in the somatic cells of both XX and XY gonads at the time of sex determination, and Hhat loss of function in mice recapitulates most of the testicular, skeletal, neuronal and growth defects observed in humans. In the developing testis, HHAT is not required for Sertoli cell commitment but plays a role in proper testis cord formation and the differentiation of fetal Leydig cells. Altogether, these results shed new light on the mechanisms of action of Hh proteins. Furthermore, they provide the first clinical evidence of the essential role played by lipid modification of Hh proteins in human testicular organogenesis and embryonic development.


Asunto(s)
Aciltransferasas/genética , Trastorno del Desarrollo Sexual 46,XY/genética , Proteínas Hedgehog/metabolismo , Lipoilación/genética , Mutación Missense , Transducción de Señal/genética , Aciltransferasas/química , Aciltransferasas/metabolismo , Secuencia de Aminoácidos , Animales , Femenino , Homocigoto , Humanos , Masculino , Ratones , Datos de Secuencia Molecular , Linaje , Homología de Secuencia de Aminoácido , Testículo/embriología
14.
Biol Reprod ; 90(2): 25, 2014 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-24352556

RESUMEN

Transcription factor GATA4 is required for the development and function of the mammalian gonads. We first reported that the GATA4 gene in both human and rodents is expressed as two major alternative transcripts that differ solely in their first untranslated exon (exon 1a vs. exon 1b). We had also showed by quantitative PCR that in mouse tissues, both Gata4 exon 1a- and 1b-containing transcripts are present in all sites that are normally positive for GATA4 protein. In adult tissues, exon 1a-containing transcripts generally predominate. A notable exception, however, is the testis where the Gata4 exon 1a and 1b transcripts exhibit a similar level of expression. We now confirm by in situ hybridization analysis that each transcript is also strongly expressed during gonad differentiation in both sexes in the rat. To gain further insights into how Gata4 gene expression is controlled, we characterized the mouse Gata4 promoter sequence located upstream of exon 1b. In vitro studies revealed that the Gata4 1b promoter is less active than the 1a promoter in several gonadal cell lines tested. Whereas we have previously shown that endogenous Gata4 transcription driven by the 1a promoter is dependent on a proximally located Ebox motif, we now show using complementary in vitro and in vivo approaches that Gata4 promoter 1b-directed expression is regulated by GATA4 itself. Thus, Gata4 transcription in the gonads and other tissues is ensured by distinct promoters that are regulated differentially and independently.


Asunto(s)
Factor de Transcripción GATA4/genética , Regulación de la Expresión Génica , Gónadas/metabolismo , Regiones Promotoras Genéticas , Animales , Células Cultivadas , Chlorocebus aethiops , Femenino , Factor de Transcripción GATA4/metabolismo , Células HeLa , Homeostasis/genética , Humanos , Masculino , Ratones , Ratas , Ratas Sprague-Dawley
15.
Open Res Eur ; 4: 68, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38883262

RESUMEN

The prevalence of hormone-related health issues caused by exposure to endocrine disrupting chemicals (EDCs) is a significant, and increasing, societal challenge. Declining fertility rates together with rising incidence rates of reproductive disorders and other endocrine-related diseases underscores the urgency in taking more action. Addressing the growing threat of EDCs in our environment demands robust and reliable test methods to assess a broad variety of endpoints relevant for endocrine disruption. EDCs also require effective regulatory frameworks, especially as the current move towards greater reliance on non-animal methods in chemical testing puts to test the current paradigm for EDC identification, which requires that an adverse effect is observed in an intact organism. Although great advances have been made in the field of predictive toxicology, disruption to the endocrine system and subsequent adverse health effects may prove particularly difficult to predict without traditional animal models. The MERLON project seeks to expedite progress by integrating multispecies molecular research, new approach methodologies (NAMs), human clinical epidemiology, and systems biology to furnish mechanistic insights and explore ways forward for NAM-based identification of EDCs. The focus is on sexual development and function, from foetal sex differentiation of the reproductive system through mini-puberty and puberty to sexual maturity. The project aims are geared towards closing existing knowledge gaps in understanding the effects of EDCs on human health to ultimately support effective regulation of EDCs in the European Union and beyond.

16.
Toxicol Sci ; 195(2): 169-183, 2023 09 28.
Artículo en Inglés | MEDLINE | ID: mdl-37505509

RESUMEN

Valproic acid (VPA) has long been the most widely used antiepileptic drug (AED) for the treatment of epilepsy, bipolar psychiatric disorders, and migraine. However, long-term VPA treatment has several adverse effects on the male reproductive system notably on endocrine functions and/or spermatic parameters. In utero exposure of the fetus to VPA is well known to be associated with a higher risk of several congenital malformations including those of male reproductive organs. Subsequent generations of AEDs, such as carbamazepine (CARB) and lamotrigine (LAM), are considered safer and are currently recommended for women of child-bearing age with epilepsy. Because anomalies of the male genital tract mostly result from endocrine imbalance during fetal life, we hypothesized that AEDs could directly impair testis differentiation. We thus aimed at identifying and characterizing the effects of VPA, CARB, and LAM on the differentiation and function of the different testicular cell types, and at understanding the mechanisms underlying these effects. By using ex vivo culture of first-trimester human fetal testes, we show that VPA induces multiple endocrine disruptive effects, compared with the milder ones caused by CARB and LAM. AED also subtly altered the germ cell lineage in distinct manners. Transcriptomic analysis of VPA-induced alterations highlighted a very broad range of effects on the fetal testis. Overall, our results show that AEDs can behave as endocrine disruptors for the human fetal testis ex vivo. This is consistent with, and likely underlies, the VPA-induced male genital tract masculinization abnormalities observed in patients.


Asunto(s)
Disruptores Endocrinos , Epilepsia , Humanos , Masculino , Femenino , Anticonvulsivantes/toxicidad , Anticonvulsivantes/uso terapéutico , Testículo , Disruptores Endocrinos/metabolismo , Ácido Valproico/toxicidad , Epilepsia/inducido químicamente , Epilepsia/tratamiento farmacológico , Epilepsia/metabolismo , Feto
17.
Genes (Basel) ; 14(2)2023 01 20.
Artículo en Inglés | MEDLINE | ID: mdl-36833200

RESUMEN

Sexual development is a complex process relying on numerous genes. Disruptions in some of these genes are known to cause differences of sexual development (DSDs). Advances in genome sequencing allowed the discovery of new genes implicated in sexual development, such as PBX1. We present here a fetus with a new PBX1 NM_002585.3: c.320G>A,p.(Arg107Gln) variant, presenting with severe DSD along with renal and lung malformations. Using CRISPR-Cas9 gene editing on HEK293T cells, we generated a KD cell line for PBX1. The KD cell line showed reduced proliferation and adhesion properties compared with HEK293T cells. HEK293T and KD cells were then transfected plasmids coding either PBX1 WT or PBX1-320G>A (mutant). WT or mutant PBX1 overexpression rescued cell proliferation in both cell lines. RNA-seq analyses showed less than 30 differentially expressed genes, in ectopic mutant-PBX1-expressing cells compared with WT-PBX1. Among them, U2AF1, encoding a splicing factor subunit, is an interesting candidate. Overall, mutant PBX1 seems to have modest effects compared with WT PBX1 in our model. However, the recurrence of PBX1 Arg107 substitution in patients with closely related phenotypes calls for its impact in human diseases. Further functional studies are needed to explore its effects on cellular metabolism.


Asunto(s)
Sistemas CRISPR-Cas , Edición Génica , Humanos , Células HEK293 , Feto , Desarrollo Sexual , Factor de Transcripción 1 de la Leucemia de Células Pre-B/genética
18.
J Biol Chem ; 286(26): 23280-95, 2011 Jul 01.
Artículo en Inglés | MEDLINE | ID: mdl-21515676

RESUMEN

Type I (α and ß) interferons (IFNs) elicit antiproliferative and antiviral activities via the surface receptor IFNAR. Serendipitous observations in transgenic mice in 1988 strongly suggested that IFNα/ß overexpression in the testis disrupts spermatogenesis. Here, we compare a new mouse strain transgenic for IFNß (Tg10) and a sister strain lacking the IFNAR1 subunit of IFNAR (Tg10-Ifnar1(-/-)), both strains expressing the transgene in the testis. The main source of IFNß RNA was the spermatid population. Importantly, the Tg10 mice, but not the double mutant Tg10-Ifnar1(-/-), showed altered spermatogenesis. The first IFNAR-dependent histological alteration was a higher apoptosis index in all germ cell categories apart from non-dividing spermatogonia. This occurred 3 weeks after the onset of IFNß production at postnatal day 20 and in the absence of somatic cell defects in terms of cell number, expression of specific cell markers, and hormonal activities. Several known interferon-stimulated genes were up-regulated in Tg10 Sertoli cells and prepachytene germ cells but not in pachytene spermatocytes and spermatids. In concordance with this, pachytene spermatocytes and spermatids isolated from wild-type testes did not display measurable amounts of IFNAR1 and phosphorylated STAT1 upon IFNß challenge in vitro, suggesting hyporesponsiveness of these cell types to IFN. At day 60, Tg10 males were sterile, and Sertoli cells showed increased amounts of anti-Mullerian hormone and decreased production of inhibin B, both probably attributable to the massive germ cell loss. Type I interferon signaling may lead to idiopathic infertilities by affecting the interplay between germ cells and Sertoli cells.


Asunto(s)
Infertilidad Masculina/metabolismo , Interferón beta/biosíntesis , Túbulos Seminíferos/metabolismo , Transducción de Señal , Espermatogénesis , Espermatozoides/metabolismo , Animales , Hormona Antimülleriana/genética , Hormona Antimülleriana/metabolismo , Apoptosis , Femenino , Infertilidad Masculina/genética , Infertilidad Masculina/patología , Inhibinas/genética , Inhibinas/metabolismo , Interferón beta/genética , Masculino , Ratones , Ratones Transgénicos , Fase Paquiteno/genética , Fosforilación/genética , Receptor de Interferón alfa y beta/genética , Receptor de Interferón alfa y beta/metabolismo , Factor de Transcripción STAT1/genética , Factor de Transcripción STAT1/metabolismo , Túbulos Seminíferos/patología , Células de Sertoli/metabolismo , Células de Sertoli/patología , Espermatozoides/patología , Factores de Tiempo
19.
Fertil Steril ; 117(6): 1279-1288, 2022 06.
Artículo en Inglés | MEDLINE | ID: mdl-35367063

RESUMEN

OBJECTIVE: To better understand the physiology of pain in pelvic pain pathological conditions, such as endometriosis, in which alterations of uterine innervation have been highlighted, we performed an anatomic and functional mapping of the macro- and microinnervation of the human uterus. Our aim was to provide a 3-dimensional reconstruction model of uterine innervation. DESIGN: This was an experimental study. We dissected the pelvises of 4 human female fetuses into serial sections, and treated them with hematoxylin and eosin staining before immunostaining. SETTING: Academic Research Unit. PATIENTS: None. INTERVENTIONS: None. MAIN OUTCOME MEASURES: Detection of nerves (S100 +) and characterization of the types of nerves. The slices obtained were aligned to construct a 3-dimensional model. RESULTS: A 3-dimensional model of uterine innervation was constructed. The nerve fibers appeared to have a centripetal path from the uterine serosa to the endometrium. Within the myometrium, innervation was dense. Endometrial innervation was sparse but present in the functional layer of the endometrium. Overall innervation was richest in the supravaginal cervix and rarer in the body of the uterus. Innervation was rich particularly laterally to the cervix next to the parametrium and paracervix. Four types of nerve fibers were identified: autonomic sympathetic (TH+), parasympathetic (VIP+), and sensitive (NPY+, CGRP1+ and VIP+). They were found in the 3 portions and the 3 layers of the uterus. CONCLUSIONS: We constructed a 3-dimensional model of the human uterine innervation. This model could provide a solid base for studying uterine innervation in pathologic situations, in order to find new therapeutic approaches.


Asunto(s)
Endometriosis , Útero , Endometriosis/patología , Endometrio/patología , Femenino , Humanos , Miometrio/patología , Dolor Pélvico/cirugía , Útero/patología
20.
J Clin Endocrinol Metab ; 107(6): 1647-1661, 2022 05 17.
Artículo en Inglés | MEDLINE | ID: mdl-35147701

RESUMEN

CONTEXT: Acetaminophen (APAP, paracetamol) is widely used by pregnant women. Although long considered safe, growing evidence indicates that APAP is an endocrine disruptor since in utero exposure may be associated with a higher risk of male genital tract abnormalities. In rodents, fetal exposure has long-term effects on the reproductive function of female offspring. Human studies have also suggested harmful APAP exposure effects. OBJECTIVE: Given that disruption of fetal ovarian development may impact women's reproductive health, we investigated the effects of APAP on fetal human ovaries in culture. DESIGN AND SETTING: Human ovarian fragments from 284 fetuses aged 7 to 12 developmental weeks (DW) were cultivated ex vivo for 7 days in the presence of human-relevant concentrations of APAP (10-8 to 10-3 M) or vehicle control. MAIN OUTCOME MEASURES: Outcomes included examination of postculture tissue morphology, cell viability, apoptosis, and quantification of hormones, APAP, and APAP metabolites in conditioned culture media. RESULTS: APAP reduced the total cell number specifically in 10- to 12-DW ovaries, induced cell death, and decreased KI67-positive cell density independently of fetal age. APAP targeted subpopulations of germ cells and disrupted human fetal ovarian steroidogenesis, without affecting prostaglandin or inhibin B production. Human fetal ovaries were able to metabolize APAP. CONCLUSIONS: Our data indicate that APAP can impact first trimester human fetal ovarian development, especially during a 10- to 12-DW window of heightened sensitivity. Overall, APAP behaves as an endocrine disruptor in the fetal human ovary.


Asunto(s)
Disruptores Endocrinos , Ovario , Acetaminofén/toxicidad , Femenino , Feto , Humanos , Masculino , Embarazo , Primer Trimestre del Embarazo
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