biostudies-arrayexpressMarie-Justine GuerquinHomo sapienshttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-13840Fetal human fetal testes (8-11 weeks post-fertilization [WPF]) were cut into small pieces (~1 mm3 ) and 4-6 pieces/testis (depending of the age or size of the gonad) were insert into the muscle of the left back of two twin female mice. One week after surgery, one mouse was exposed by drinking water to a mixture of BPA (1 µM, CAS no. 80-09-1, Merck) and DEHP (1 µM, CAS no. 117-81-7, Merck) and the other was exposed by drinking water to the vehicle (absolute ethanol [ETOH] diluted at 1/1000000 in water). Mice were exposed during 6-11 weeks (corresponding to the 18 WPF equivalent age) and at the end of exposure, testes were collected and each pieces were cleaned from extracellular matrix and murine muscular tissue. One part of the pieces were used for RRBS analyses.biostudies-arrayexpressLibrary Construction - RRBS was performed by Integragen SA (Evry, France) with the Diagenode Premium RRBS kit. In brief, 100 ng of qualified genomic DNA were digested with MspI. After end-repair, A-tailing, and ligation to methylated and indexed adapters, the size selected library fragments were subjected to bisulfite conversion and PCR amplified.Sequencing - amplified DNA was sequenced on an Illumina HiSeq4000 sequencer as Paired_End 75 bp reads. Image analysis and base calling is performed using Illumina Real Time Analysis (2.7.7) with default parameters. Base calling is performed by using the Real-Time Analysis software sequence pipeline (2.7.7) with default parameters.Nucleic Acid Extraction - Total DNA from xenografted testes were simultaneously isolated using the AllPrep DNA/RNA microkit (ref : 80284, Qiagen, France) according to manufacturer’s instructions. DNA concentrations and quality were assayed with Qbit fluorometer (Thermofisher) and Agilent bioanalyser. After DNA quantitation, pools of DNA per condition were performed containing from 3 to 8 testis per pool.Sample Treatment - Xenografted CTL mice (control) were orally exposed (drinking water) to vehicle (ETOH, 1/1000000) and DE/BP mice were exposed to a mixture of DEHP and BPA at 1µM each.Sample Collection - At the end of exposure, fetal human testis were collected and each pieces were cleaned from extracellular matrix and murine muscular tissue. Then, testicular pieces were pooled by condition and stored in RLT buffer from AllPrep DNA/RNA microkit (Qiagen).Growth Protocol - human fetal testis (from 8-11 WPF) were cut into small pieces (~1 mm3 ) and 4-6 pieces/testis (depending of the age or size of the gonad) were insert into the muscle of the left back of two twin female mice. One week after surgery, mice were orally exposed to chemicals during 6-11 weeks (corresponding to the 18 WPF equivalent age)OrganizationMINSEQE ScoreAssays and DataProcessed DataMAGE-TAB FilesData Transformation - Differential analysis was perform with methylKit R package to identify DMC (differential methylated CpG) or DMR (differentiel methylated regions of CpG) that were covered by at least 10 reads in all samples. DMR were defined with parameters : win.size=300 and step.size=300 . DMC and DMR were considered as significant with a delta of percentage of methylation > 25 % and an adjusted p.value < 0.05.Sequence Alignment - We used BS-Seeker2 (v2.1.8) to map RRBS data to the human genome (hg38) and retrieve the number of methylated and unmethylated cytosines at each covered CpG site. Allowing local/gapped alignment with Bowtie2, increased the mappability. The parameters used are: -r (Map reads to the Reduced Representation genome), -c C-CGG (MspI: sites of restriction enzyme and specifying lengths of fragments ranging [40bp, 400bp]. One mismatch is allowed in adaptor sequence.UnknownTranscriptomicsGenomicsProteomicsIllumina HiSeq 4000<h4>ABSTRACT</h4> Testicular cancer is an increasing burden in modern societies and the most common malignancy among young adult males. Environment contaminants, especially endocrine disrupting compounds (EDC), may play a significant role in the development of these cancers through epigenetic alterations occurring during fetal and neonatal development. As long-term studies in humans and suitable experimental models with the potential to develop testicular cancer are lacking, no causal link can be established between endocrine disruptor exposure and testicular cancer incidence. Therefore, we developed an experimental model that recapitulates the differentiation of germ cells from primordial germ cells (pluripotency) into spermatocytes (meiosis) by using xenografted human fetal testis combined with germ cell transplantation into adult testis compartments. Using this model, we demonstrate that long-term fetal exposure (until 12 weeks) to a mixture of Di-2-ethylhexylphthalate (DEHP) and Bisphenol A (BPA), two most prevalent plasticizers, could interfere with fetal germ cell differentiation, leading to carcinogenesis and seminomas. Transcriptome, methylome, and histological analyses reveal that BPA/DEHP exposure induced some significant hallmarks of germ cell tumors to occur: persistent pluripotent and proliferative germ cells, global hypomethylation of CpGs in germ cells, abnormal expression of meiotic markers and fibrotic signatures in fetal testis. Additionally, we found that EDC-exposed fetal germ cells were more likely to develop seminoma in a context that allows spermatogenesis to begin. This study proposes the first experimental evidence that EDC exposure can cause long-term, irreversible lesions in fetal germ cells, which then lead to testicular tumorigenesis in adults.methylation profiling by high throughput sequencingHomo sapiensEvidence of the ability of endocrine disrupting compounds to induce testicular germ cell cancer in humansNour Nicolas, Delphine Moison, Amandine Jampy, Quentin Masson, Nathalie Dechamps, Sébastien Messiaen, Sonia Abdallah, Stéphanie Pozzi-Gaudin, Alexandra Benachi, Damien Ulveling, Claire Francastel, Virginie Rouiller-Fabre, Gabriel Livera, Marie-Justine GuerquinMarie-Justine GuerquinfalseRRBS of human fetal testis treated to a mixture of DEHP/BPAFetal human fetal testes (8-11 weeks post-fertilization [WPF]) were cut into small pieces (~1 mm3 ) and 4-6 pieces/testis (depending of the age or size of the gonad) were insert into the muscle of the left back of two twin female mice. One week after surgery, one mouse was exposed by drinking water to a mixture of BPA (1 µM, CAS no. 80-09-1, Merck) and DEHP (1 µM, CAS no. 117-81-7, Merck) and the other was exposed by drinking water to the vehicle (absolute ethanol [ETOH] diluted at 1/1000000 in water). Mice were exposed during 6-11 weeks (corresponding to the 18 WPF equivalent age) and at the end of exposure, testes were collected and each pieces were cleaned from extracellular matrix and murine muscular tissue. One part of the pieces were used for RRBS analyses.2026-01-01T00:00:00Z2026-01-01T02:02:01.552Z2024-02-20T23:07:50.909ZE-MTAB-13840ERP157632E-MTAB-13835EFO_0002944EFO_0004170EFO_0003789EFO_0002761EFO_0004917EFO_0005518EFO_0003816EFO_0004184EFO_000396910.1101/2023.12.22.573063