WP1: To characterize the cellular composition of the human foetal gonads, we have analysed the DNA, RNA or protein content of over 1,4 million single cells from 230 human embryos/foetuses aged between 5 to 21 post-conception weeks. Our results allowed us to identify known and new cell types and to dissect the molecular mechanisms involved in the early specification of gonads into either an ovary or a testis. In addition, we could highlight specific genes and proteins that can be used as markers to map these cells inside the gonads. Several of these genes could represent new biomarkers for DSDs.
WP2: We have generated spatially resolved transcriptomic profiling data from human developmental gonads at different time points, and using three orthogonal technologies: Nanostring WTA, 10xGenomics Visium, and 10xGenomics Cartana. This data will combined with scRNA sequencing dat,a provide a much more detailed view over the spatiotemporal development of the cellular diversity of this organ during the studied time window.
WP3: A total of 156 human foetal gonads were used to assess the spatial location of gonadal cells in situ using state-of-the-art techniques implemented in WP3. We performed, fully automated cyclic immuno-fluorescence imaging of frozen sections of gonads. We also imaged in 3D, solvent-cleared human gonads, genital ducts and genitalia using light sheet fluorescence microscopy (LSFM) and fluorescent in situ hybridization. We have been able to describe the cellular composition and organization of the developing human gonads.
WP4: Keen Eye completed the deliverable for D4.2 which is an enhanced 3D viewer for image data. With respect to D4.3 and D4.4 Keen Eye rebuilt large parts of the annotation rendering engine (for handling up to 10M Annotations), performed an investigation into 3D registration algorithms and techniques, and refactored the platform to allow such registration algorithms to run. The final integration and testing on the platform was not completed.
WP5: A total of 273 human fetal gonads were cultured ex vivo and signaling were manipulated to mimic conditions of Differences of Sex Development or effects of endocrine disruptors. The induced alterations were examined at molecular and cellular levels by established endpoints in combination with state-of-the-art techniques implemented in WPs 1-4. The inclusion of novel techniques significantly improved the analysis and thus the relevance of the obtained results.
WP6: P01a and P09 have organized in Paris a tissue-clearing and 3D imaging course freely accessible to members of teams belonging to the cluster of 6 H2020/HCA consortia. A spatial transcriptomics course was organized (with EMBO support) in Solna (Sweden) by P02 and P03.We have also organized in Paris a one-day final public symposium with international speakers. A video and a short animation, describing in lay language the objectives and partners of HUGODECA were produced and released on social media.
WP7: The goal of WP7 was to establish some guidelines about the management of the data collected, generated or processed during the HUGODECA project, so as to make them Findable, Accessible, Interoperable and Re-usable (FAIR principles) by the scientific community. We defined those guidelines in the Data Management Plan, we set up a platform to share metadata templates and we took part in the establishment of pipelines and dataset preparation.
