--- _id: '15048' abstract: - lang: eng text: Embryogenesis results from the coordinated activities of different signaling pathways controlling cell fate specification and morphogenesis. In vertebrate gastrulation, both Nodal and BMP signaling play key roles in germ layer specification and morphogenesis, yet their interplay to coordinate embryo patterning with morphogenesis is still insufficiently understood. Here, we took a reductionist approach using zebrafish embryonic explants to study the coordination of Nodal and BMP signaling for embryo patterning and morphogenesis. We show that Nodal signaling triggers explant elongation by inducing mesendodermal progenitors but also suppressing BMP signaling activity at the site of mesendoderm induction. Consistent with this, ectopic BMP signaling in the mesendoderm blocks cell alignment and oriented mesendoderm intercalations, key processes during explant elongation. Translating these ex vivo observations to the intact embryo showed that, similar to explants, Nodal signaling suppresses the effect of BMP signaling on cell intercalations in the dorsal domain, thus allowing robust embryonic axis elongation. These findings suggest a dual function of Nodal signaling in embryonic axis elongation by both inducing mesendoderm and suppressing BMP effects in the dorsal portion of the mesendoderm. acknowledged_ssus: - _id: Bio - _id: LifeSc acknowledgement: "We thank Patrick Müller for sharing the chordintt250 mutant zebrafish line as well as the plasmid for chrd-GFP, Katherine Rogers for sharing the bmp2b plasmid and Andrea Pauli for sharing the draculin plasmid. Diana Pinheiro generated the MZlefty1,2;Tg(sebox::EGFP) line. We are grateful to Patrick Müller, Diana Pinheiro and Katherine Rogers and members of the Heisenberg lab for discussions, technical advice and feedback on the manuscript. We also thank Anna Kicheva and Edouard Hannezo for discussions. We thank the Imaging and Optics Facility as well as the Life Science facility at IST Austria for support with microscopy and fish maintenance.\r\nThis work was supported by a European Research Council Advanced Grant\r\n(MECSPEC 742573 to C.-P.H.). A.S. is a recipient of a DOC Fellowship of the Austrian\r\nAcademy of Sciences at IST Austria. Open Access funding provided by Institute of\r\nScience and Technology Austria. " article_processing_charge: Yes (via OA deal) article_type: original author: - first_name: Alexandra full_name: Schauer, Alexandra id: 30A536BA-F248-11E8-B48F-1D18A9856A87 last_name: Schauer orcid: 0000-0001-7659-9142 - first_name: Kornelija full_name: Pranjic-Ferscha, Kornelija id: 4362B3C2-F248-11E8-B48F-1D18A9856A87 last_name: Pranjic-Ferscha - first_name: Robert full_name: Hauschild, Robert id: 4E01D6B4-F248-11E8-B48F-1D18A9856A87 last_name: Hauschild orcid: 0000-0001-9843-3522 - first_name: Carl-Philipp J full_name: Heisenberg, Carl-Philipp J id: 39427864-F248-11E8-B48F-1D18A9856A87 last_name: Heisenberg orcid: 0000-0002-0912-4566 citation: ama: Schauer A, Pranjic-Ferscha K, Hauschild R, Heisenberg C-PJ. Robust axis elongation by Nodal-dependent restriction of BMP signaling. Development. 2024;151(4):1-18. doi:10.1242/dev.202316 apa: Schauer, A., Pranjic-Ferscha, K., Hauschild, R., & Heisenberg, C.-P. J. (2024). Robust axis elongation by Nodal-dependent restriction of BMP signaling. Development. The Company of Biologists. https://doi.org/10.1242/dev.202316 chicago: Schauer, Alexandra, Kornelija Pranjic-Ferscha, Robert Hauschild, and Carl-Philipp J Heisenberg. “Robust Axis Elongation by Nodal-Dependent Restriction of BMP Signaling.” Development. The Company of Biologists, 2024. https://doi.org/10.1242/dev.202316. ieee: A. Schauer, K. Pranjic-Ferscha, R. Hauschild, and C.-P. J. Heisenberg, “Robust axis elongation by Nodal-dependent restriction of BMP signaling,” Development, vol. 151, no. 4. The Company of Biologists, pp. 1–18, 2024. ista: Schauer A, Pranjic-Ferscha K, Hauschild R, Heisenberg C-PJ. 2024. Robust axis elongation by Nodal-dependent restriction of BMP signaling. Development. 151(4), 1–18. mla: Schauer, Alexandra, et al. “Robust Axis Elongation by Nodal-Dependent Restriction of BMP Signaling.” Development, vol. 151, no. 4, The Company of Biologists, 2024, pp. 1–18, doi:10.1242/dev.202316. short: A. Schauer, K. Pranjic-Ferscha, R. Hauschild, C.-P.J. Heisenberg, Development 151 (2024) 1–18. date_created: 2024-03-03T23:00:50Z date_published: 2024-02-01T00:00:00Z date_updated: 2024-03-04T07:28:25Z day: '01' ddc: - '570' department: - _id: CaHe - _id: Bio doi: 10.1242/dev.202316 ec_funded: 1 file: - access_level: open_access checksum: 6961ea10012bf0d266681f9628bb8f13 content_type: application/pdf creator: dernst date_created: 2024-03-04T07:24:43Z date_updated: 2024-03-04T07:24:43Z file_id: '15050' file_name: 2024_Development_Schauer.pdf file_size: 14839986 relation: main_file success: 1 file_date_updated: 2024-03-04T07:24:43Z has_accepted_license: '1' intvolume: ' 151' issue: '4' language: - iso: eng license: https://creativecommons.org/licenses/by/4.0/ month: '02' oa: 1 oa_version: Published Version page: 1-18 project: - _id: 260F1432-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '742573' name: Interaction and feedback between cell mechanics and fate specification in vertebrate gastrulation - _id: 26B1E39C-B435-11E9-9278-68D0E5697425 grant_number: '25239' name: 'Mesendoderm specification in zebrafish: The role of extraembryonic tissues' publication: Development publication_identifier: eissn: - 1477-9129 issn: - 0950-1991 publication_status: published publisher: The Company of Biologists quality_controlled: '1' related_material: record: - id: '14926' relation: research_data status: public scopus_import: '1' status: public title: Robust axis elongation by Nodal-dependent restriction of BMP signaling tmp: image: /images/cc_by.png legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0) short: CC BY (4.0) type: journal_article user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87 volume: 151 year: '2024' ... --- _id: '14926' author: - first_name: Robert full_name: Hauschild, Robert id: 4E01D6B4-F248-11E8-B48F-1D18A9856A87 last_name: Hauschild orcid: 0000-0001-9843-3522 citation: ama: Hauschild R. Matlab script for analysis of clone dispersal. 2024. doi:10.15479/AT:ISTA:14926 apa: Hauschild, R. (2024). Matlab script for analysis of clone dispersal. ISTA. https://doi.org/10.15479/AT:ISTA:14926 chicago: Hauschild, Robert. “Matlab Script for Analysis of Clone Dispersal.” ISTA, 2024. https://doi.org/10.15479/AT:ISTA:14926. ieee: R. Hauschild, “Matlab script for analysis of clone dispersal.” ISTA, 2024. ista: Hauschild R. 2024. Matlab script for analysis of clone dispersal, ISTA, 10.15479/AT:ISTA:14926. mla: Hauschild, Robert. Matlab Script for Analysis of Clone Dispersal. ISTA, 2024, doi:10.15479/AT:ISTA:14926. short: R. Hauschild, (2024). date_created: 2024-02-02T14:42:26Z date_published: 2024-02-02T00:00:00Z date_updated: 2024-03-04T07:28:25Z day: '02' ddc: - '570' department: - _id: Bio doi: 10.15479/AT:ISTA:14926 file: - access_level: open_access checksum: df7f358ae19a176cf710c0a802ce31b1 content_type: application/octet-stream creator: rhauschild date_created: 2024-02-02T14:40:31Z date_updated: 2024-02-02T14:40:31Z file_id: '14927' file_name: README.md file_size: 736 relation: main_file success: 1 - access_level: open_access checksum: 10194cc11619eccd8f4b24472e465b7f content_type: application/x-zip-compressed creator: rhauschild date_created: 2024-02-02T14:40:31Z date_updated: 2024-02-02T14:40:31Z file_id: '14928' file_name: Supplementary_file_1.zip file_size: 3543 relation: main_file success: 1 file_date_updated: 2024-02-02T14:40:31Z has_accepted_license: '1' license: https://opensource.org/licenses/MIT month: '02' oa: 1 publisher: ISTA related_material: record: - id: '15048' relation: used_in_publication status: public status: public title: Matlab script for analysis of clone dispersal tmp: legal_code_url: https://opensource.org/licenses/MIT name: The MIT License short: MIT type: software user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87 year: '2024' ... --- _id: '15146' abstract: - lang: eng text: The extracellular matrix (ECM) serves as a scaffold for cells and plays an essential role in regulating numerous cellular processes, including cell migration and proliferation. Due to limitations in specimen preparation for conventional room-temperature electron microscopy, we lack structural knowledge on how ECM components are secreted, remodeled, and interact with surrounding cells. We have developed a 3D-ECM platform compatible with sample thinning by cryo-focused ion beam milling, the lift-out extraction procedure, and cryo-electron tomography. Our workflow implements cell-derived matrices (CDMs) grown on EM grids, resulting in a versatile tool closely mimicking ECM environments. This allows us to visualize ECM for the first time in its hydrated, native context. Our data reveal an intricate network of extracellular fibers, their positioning relative to matrix-secreting cells, and previously unresolved structural entities. Our workflow and results add to the structural atlas of the ECM, providing novel insights into its secretion and assembly. acknowledged_ssus: - _id: LifeSc - _id: ScienComp - _id: EM-Fac - _id: M-Shop acknowledgement: "Open Access funding provided by IST Austria. We thank Armel Nicolas and his team at the ISTA proteomics facility, Alois Schloegl, Stefano Elefante, and colleagues at the ISTA Scientific Computing facility, Tommaso Constanzo and Ludek Lovicar at the Electron Microsocpy Facility (EMF), and Thomas Menner at the Miba Machine shop for their support. We also thank Wanda Kukulski (University of Bern) as well as Darío Porley, Andreas Thader, and other members of the Schur group for helpful discussions. Matt Swulius and Jessica Heebner provided great support in using Dragonfly. We thank Dorotea Fracciolla (Art & Science) for support in figure illustration.\r\n\r\nThis research was supported by the Scientific Service Units of ISTA through resources provided by Scientific Computing, the Lab Support Facility, and the Electron Microscopy Facility. We acknowledge funding support from the following sources: Austrian Science Fund (FWF) grant P33367 (to F.K.M. Schur), the Federation of European Biochemical Societies (to F.K.M. Schur), Niederösterreich (NÖ) Fonds (to B. Zens), FWF grant E435 (to J.M. Hansen), European Research Council under the European Union’s Horizon 2020 research (grant agreement No. 724373) (to M. Sixt), and Jenny and Antti Wihuri Foundation (to J. Alanko). This publication has been made possible in part by CZI grant DAF2021-234754 and grant DOI https://doi.org/10.37921/812628ebpcwg from the Chan Zuckerberg Initiative DAF, an advised fund of Silicon Valley Community Foundation (to F.K.M. Schur)." article_number: e202309125 article_processing_charge: Yes (via OA deal) article_type: original author: - first_name: Bettina full_name: Zens, Bettina id: 45FD126C-F248-11E8-B48F-1D18A9856A87 last_name: Zens - first_name: Florian full_name: Fäßler, Florian id: 404F5528-F248-11E8-B48F-1D18A9856A87 last_name: Fäßler orcid: 0000-0001-7149-769X - first_name: Jesse full_name: Hansen, Jesse id: 1063c618-6f9b-11ec-9123-f912fccded63 last_name: Hansen - first_name: Robert full_name: Hauschild, Robert id: 4E01D6B4-F248-11E8-B48F-1D18A9856A87 last_name: Hauschild orcid: 0000-0001-9843-3522 - first_name: Julia full_name: Datler, Julia id: 3B12E2E6-F248-11E8-B48F-1D18A9856A87 last_name: Datler orcid: 0000-0002-3616-8580 - first_name: Victor-Valentin full_name: Hodirnau, Victor-Valentin id: 3661B498-F248-11E8-B48F-1D18A9856A87 last_name: Hodirnau - first_name: Vanessa full_name: Zheden, Vanessa id: 39C5A68A-F248-11E8-B48F-1D18A9856A87 last_name: Zheden orcid: 0000-0002-9438-4783 - first_name: Jonna H full_name: Alanko, Jonna H id: 2CC12E8C-F248-11E8-B48F-1D18A9856A87 last_name: Alanko orcid: 0000-0002-7698-3061 - first_name: Michael K full_name: Sixt, Michael K id: 41E9FBEA-F248-11E8-B48F-1D18A9856A87 last_name: Sixt orcid: 0000-0002-6620-9179 - first_name: Florian KM full_name: Schur, Florian KM id: 48AD8942-F248-11E8-B48F-1D18A9856A87 last_name: Schur orcid: 0000-0003-4790-8078 citation: ama: Zens B, Fäßler F, Hansen J, et al. Lift-out cryo-FIBSEM and cryo-ET reveal the ultrastructural landscape of extracellular matrix. Journal of Cell Biology. 2024;223(6). doi:10.1083/jcb.202309125 apa: Zens, B., Fäßler, F., Hansen, J., Hauschild, R., Datler, J., Hodirnau, V.-V., … Schur, F. K. (2024). Lift-out cryo-FIBSEM and cryo-ET reveal the ultrastructural landscape of extracellular matrix. Journal of Cell Biology. Rockefeller University Press. https://doi.org/10.1083/jcb.202309125 chicago: Zens, Bettina, Florian Fäßler, Jesse Hansen, Robert Hauschild, Julia Datler, Victor-Valentin Hodirnau, Vanessa Zheden, Jonna H Alanko, Michael K Sixt, and Florian KM Schur. “Lift-out Cryo-FIBSEM and Cryo-ET Reveal the Ultrastructural Landscape of Extracellular Matrix.” Journal of Cell Biology. Rockefeller University Press, 2024. https://doi.org/10.1083/jcb.202309125. ieee: B. Zens et al., “Lift-out cryo-FIBSEM and cryo-ET reveal the ultrastructural landscape of extracellular matrix,” Journal of Cell Biology, vol. 223, no. 6. Rockefeller University Press, 2024. ista: Zens B, Fäßler F, Hansen J, Hauschild R, Datler J, Hodirnau V-V, Zheden V, Alanko JH, Sixt MK, Schur FK. 2024. Lift-out cryo-FIBSEM and cryo-ET reveal the ultrastructural landscape of extracellular matrix. Journal of Cell Biology. 223(6), e202309125. mla: Zens, Bettina, et al. “Lift-out Cryo-FIBSEM and Cryo-ET Reveal the Ultrastructural Landscape of Extracellular Matrix.” Journal of Cell Biology, vol. 223, no. 6, e202309125, Rockefeller University Press, 2024, doi:10.1083/jcb.202309125. short: B. Zens, F. Fäßler, J. Hansen, R. Hauschild, J. Datler, V.-V. Hodirnau, V. Zheden, J.H. Alanko, M.K. Sixt, F.K. Schur, Journal of Cell Biology 223 (2024). date_created: 2024-03-21T06:45:51Z date_published: 2024-03-20T00:00:00Z date_updated: 2024-03-25T13:03:57Z day: '20' ddc: - '570' department: - _id: FlSc - _id: MiSi - _id: Bio - _id: EM-Fac doi: 10.1083/jcb.202309125 ec_funded: 1 external_id: pmid: - '38506714' file: - access_level: open_access checksum: 90d1984a93660735e506c2a304bc3f73 content_type: application/pdf creator: dernst date_created: 2024-03-25T12:52:04Z date_updated: 2024-03-25T12:52:04Z file_id: '15188' file_name: 2024_JCB_Zens.pdf file_size: 11907016 relation: main_file success: 1 file_date_updated: 2024-03-25T12:52:04Z has_accepted_license: '1' intvolume: ' 223' issue: '6' language: - iso: eng month: '03' oa: 1 oa_version: Published Version pmid: 1 project: - _id: 9B954C5C-BA93-11EA-9121-9846C619BF3A grant_number: P33367 name: Structure and isoform diversity of the Arp2/3 complex - _id: 7bd318a1-9f16-11ee-852c-cc9217763180 grant_number: E435 name: In Situ Actin Structures via Hybrid Cryo-electron Microscopy - _id: 25FE9508-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '724373' name: Cellular navigation along spatial gradients - _id: 059B463C-7A3F-11EA-A408-12923DDC885E name: NÖ-Fonds Preis für die Jungforscherin des Jahres am IST Austria - _id: 2615199A-B435-11E9-9278-68D0E5697425 grant_number: '21317' name: Spatiotemporal regulation of chemokine-induced signalling in leukocyte chemotaxis - _id: 62909c6f-2b32-11ec-9570-e1476aab5308 grant_number: CZI01 name: CryoMinflux-guided in-situ visual proteomics and structure determination publication: Journal of Cell Biology publication_identifier: eissn: - 1540-8140 issn: - 0021-9525 publication_status: published publisher: Rockefeller University Press quality_controlled: '1' scopus_import: '1' status: public title: Lift-out cryo-FIBSEM and cryo-ET reveal the ultrastructural landscape of extracellular matrix tmp: image: /images/cc_by.png legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0) short: CC BY (4.0) type: journal_article user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87 volume: 223 year: '2024' ... --- _id: '12830' abstract: - lang: eng text: Interstitial fluid (IF) accumulation between embryonic cells is thought to be important for embryo patterning and morphogenesis. Here, we identify a positive mechanical feedback loop between cell migration and IF relocalization and find that it promotes embryonic axis formation during zebrafish gastrulation. We show that anterior axial mesendoderm (prechordal plate [ppl]) cells, moving in between the yolk cell and deep cell tissue to extend the embryonic axis, compress the overlying deep cell layer, thereby causing IF to flow from the deep cell layer to the boundary between the yolk cell and the deep cell layer, directly ahead of the advancing ppl. This IF relocalization, in turn, facilitates ppl cell protrusion formation and migration by opening up the space into which the ppl moves and, thereby, the ability of the ppl to trigger IF relocalization by pushing against the overlying deep cell layer. Thus, embryonic axis formation relies on a hydraulic feedback loop between cell migration and IF relocalization. acknowledged_ssus: - _id: PreCl - _id: Bio acknowledgement: We thank Andrea Pauli (IMP) and Edouard Hannezo (ISTA) for fruitful discussions and support with the SPIM experiments; the Heisenberg group, and especially Feyza Nur Arslan and Alexandra Schauer, for discussions and feedback; Michaela Jović (ISTA) for help with the quantitative real-time PCR protocol; the bioimaging and zebrafish facilities of ISTA for continuous support; Stephan Preibisch (Janelia Research Campus) for support with the SPIM data analysis; and Nobuhiro Nakamura (Tokyo Institute of Technology) for sharing α1-Na+/K+-ATPase antibody. This work was supported by funding from the European Union (European Research Council Advanced grant 742573 to C.-P.H.), postdoctoral fellowships from EMBO (LTF-850-2017) and HFSP (LT000429/2018-L2) to D.P., and a PhD fellowship from the Studienstiftung des deutschen Volkes to F.P. article_processing_charge: Yes (via OA deal) article_type: original author: - first_name: Karla full_name: Huljev, Karla id: 44C6F6A6-F248-11E8-B48F-1D18A9856A87 last_name: Huljev - first_name: Shayan full_name: Shamipour, Shayan id: 40B34FE2-F248-11E8-B48F-1D18A9856A87 last_name: Shamipour - first_name: Diana C full_name: Nunes Pinheiro, Diana C id: 2E839F16-F248-11E8-B48F-1D18A9856A87 last_name: Nunes Pinheiro orcid: 0000-0003-4333-7503 - first_name: Friedrich full_name: Preusser, Friedrich last_name: Preusser - first_name: Irene full_name: Steccari, Irene id: 2705C766-9FE2-11EA-B224-C6773DDC885E last_name: Steccari - first_name: Christoph M full_name: Sommer, Christoph M id: 4DF26D8C-F248-11E8-B48F-1D18A9856A87 last_name: Sommer orcid: 0000-0003-1216-9105 - first_name: Suyash full_name: Naik, Suyash id: 2C0B105C-F248-11E8-B48F-1D18A9856A87 last_name: Naik orcid: 0000-0001-8421-5508 - first_name: Carl-Philipp J full_name: Heisenberg, Carl-Philipp J id: 39427864-F248-11E8-B48F-1D18A9856A87 last_name: Heisenberg orcid: 0000-0002-0912-4566 citation: ama: Huljev K, Shamipour S, Nunes Pinheiro DC, et al. A hydraulic feedback loop between mesendoderm cell migration and interstitial fluid relocalization promotes embryonic axis formation in zebrafish. Developmental Cell. 2023;58(7):582-596.e7. doi:10.1016/j.devcel.2023.02.016 apa: Huljev, K., Shamipour, S., Nunes Pinheiro, D. C., Preusser, F., Steccari, I., Sommer, C. M., … Heisenberg, C.-P. J. (2023). A hydraulic feedback loop between mesendoderm cell migration and interstitial fluid relocalization promotes embryonic axis formation in zebrafish. Developmental Cell. Elsevier. https://doi.org/10.1016/j.devcel.2023.02.016 chicago: Huljev, Karla, Shayan Shamipour, Diana C Nunes Pinheiro, Friedrich Preusser, Irene Steccari, Christoph M Sommer, Suyash Naik, and Carl-Philipp J Heisenberg. “A Hydraulic Feedback Loop between Mesendoderm Cell Migration and Interstitial Fluid Relocalization Promotes Embryonic Axis Formation in Zebrafish.” Developmental Cell. Elsevier, 2023. https://doi.org/10.1016/j.devcel.2023.02.016. ieee: K. Huljev et al., “A hydraulic feedback loop between mesendoderm cell migration and interstitial fluid relocalization promotes embryonic axis formation in zebrafish,” Developmental Cell, vol. 58, no. 7. Elsevier, p. 582–596.e7, 2023. ista: Huljev K, Shamipour S, Nunes Pinheiro DC, Preusser F, Steccari I, Sommer CM, Naik S, Heisenberg C-PJ. 2023. A hydraulic feedback loop between mesendoderm cell migration and interstitial fluid relocalization promotes embryonic axis formation in zebrafish. Developmental Cell. 58(7), 582–596.e7. mla: Huljev, Karla, et al. “A Hydraulic Feedback Loop between Mesendoderm Cell Migration and Interstitial Fluid Relocalization Promotes Embryonic Axis Formation in Zebrafish.” Developmental Cell, vol. 58, no. 7, Elsevier, 2023, p. 582–596.e7, doi:10.1016/j.devcel.2023.02.016. short: K. Huljev, S. Shamipour, D.C. Nunes Pinheiro, F. Preusser, I. Steccari, C.M. Sommer, S. Naik, C.-P.J. Heisenberg, Developmental Cell 58 (2023) 582–596.e7. date_created: 2023-04-16T22:01:07Z date_published: 2023-04-10T00:00:00Z date_updated: 2023-08-01T14:10:38Z day: '10' ddc: - '570' department: - _id: CaHe - _id: Bio doi: 10.1016/j.devcel.2023.02.016 ec_funded: 1 external_id: isi: - '000982111800001' file: - access_level: open_access checksum: c80ca2ebc241232aacdb5aa4b4c80957 content_type: application/pdf creator: dernst date_created: 2023-04-17T07:41:25Z date_updated: 2023-04-17T07:41:25Z file_id: '12842' file_name: 2023_DevelopmentalCell_Huljev.pdf file_size: 7925886 relation: main_file success: 1 file_date_updated: 2023-04-17T07:41:25Z has_accepted_license: '1' intvolume: ' 58' isi: 1 issue: '7' language: - iso: eng month: '04' oa: 1 oa_version: Published Version page: 582-596.e7 project: - _id: 260F1432-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '742573' name: Interaction and feedback between cell mechanics and fate specification in vertebrate gastrulation - _id: 26520D1E-B435-11E9-9278-68D0E5697425 grant_number: ALTF 850-2017 name: Coordination of mesendoderm cell fate specification and internalization during zebrafish gastrulation - _id: 266BC5CE-B435-11E9-9278-68D0E5697425 grant_number: LT000429 name: Coordination of mesendoderm fate specification and internalization during zebrafish gastrulation publication: Developmental Cell publication_identifier: eissn: - 1878-1551 issn: - 1534-5807 publication_status: published publisher: Elsevier quality_controlled: '1' scopus_import: '1' status: public title: A hydraulic feedback loop between mesendoderm cell migration and interstitial fluid relocalization promotes embryonic axis formation in zebrafish tmp: image: /images/cc_by.png legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0) short: CC BY (4.0) type: journal_article user_id: 4359f0d1-fa6c-11eb-b949-802e58b17ae8 volume: 58 year: '2023' ... --- _id: '13033' abstract: - lang: eng text: Current methods for assessing cell proliferation in 3D scaffolds rely on changes in metabolic activity or total DNA, however, direct quantification of cell number in 3D scaffolds remains a challenge. To address this issue, we developed an unbiased stereology approach that uses systematic-random sampling and thin focal-plane optical sectioning of the scaffolds followed by estimation of total cell number (StereoCount). This approach was validated against an indirect method for measuring the total DNA (DNA content); and the Bürker counting chamber, the current reference method for quantifying cell number. We assessed the total cell number for cell seeding density (cells per unit volume) across four values and compared the methods in terms of accuracy, ease-of-use and time demands. The accuracy of StereoCount markedly outperformed the DNA content for cases with ~ 10,000 and ~ 125,000 cells/scaffold. For cases with ~ 250,000 and ~ 375,000 cells/scaffold both StereoCount and DNA content showed lower accuracy than the Bürker but did not differ from each other. In terms of ease-of-use, there was a strong advantage for the StereoCount due to output in terms of absolute cell numbers along with the possibility for an overview of cell distribution and future use of automation for high throughput analysis. Taking together, the StereoCount method is an efficient approach for direct cell quantification in 3D collagen scaffolds. Its major benefit is that automated StereoCount could accelerate research using 3D scaffolds focused on drug discovery for a wide variety of human diseases. acknowledgement: The study was supported by Project No. CZ.02.1.01/0.0/0.0/16_019/0000787 “Fighting INfectious Diseases”, awarded by the MEYS CR, financed from EFRR, by the Cooperatio Program, research area DIAG and research area MED/DIAG, by the profiBONE project (TO01000309) benefitting from a € (1.433.000) grant from Iceland, Liechtenstein and Norway through the EEA Grants and the Technology Agency of the Czech Republic and by a Grant (#1926990) to PRM and SRC Biosciences from the National Science Foundation (U.S. Public Health Service). The authors acknowledge the invaluable assistance provided by Iveta Paurova via her support in terms of the provision of laboratory services. article_number: '7959' article_processing_charge: No article_type: original author: - first_name: Anna full_name: Zavadakova, Anna last_name: Zavadakova - first_name: Lucie full_name: Vistejnova, Lucie last_name: Vistejnova - first_name: Tereza full_name: Belinova, Tereza id: 0bf89b6a-d28b-11eb-8bd6-f43768e4d368 last_name: Belinova - first_name: Filip full_name: Tichanek, Filip last_name: Tichanek - first_name: Dagmar full_name: Bilikova, Dagmar last_name: Bilikova - first_name: Peter R. full_name: Mouton, Peter R. last_name: Mouton citation: ama: Zavadakova A, Vistejnova L, Belinova T, Tichanek F, Bilikova D, Mouton PR. Novel stereological method for estimation of cell counts in 3D collagen scaffolds. Scientific Reports. 2023;13(1). doi:10.1038/s41598-023-35162-z apa: Zavadakova, A., Vistejnova, L., Belinova, T., Tichanek, F., Bilikova, D., & Mouton, P. R. (2023). Novel stereological method for estimation of cell counts in 3D collagen scaffolds. Scientific Reports. Springer Nature. https://doi.org/10.1038/s41598-023-35162-z chicago: Zavadakova, Anna, Lucie Vistejnova, Tereza Belinova, Filip Tichanek, Dagmar Bilikova, and Peter R. Mouton. “Novel Stereological Method for Estimation of Cell Counts in 3D Collagen Scaffolds.” Scientific Reports. Springer Nature, 2023. https://doi.org/10.1038/s41598-023-35162-z. ieee: A. Zavadakova, L. Vistejnova, T. Belinova, F. Tichanek, D. Bilikova, and P. R. Mouton, “Novel stereological method for estimation of cell counts in 3D collagen scaffolds,” Scientific Reports, vol. 13, no. 1. Springer Nature, 2023. ista: Zavadakova A, Vistejnova L, Belinova T, Tichanek F, Bilikova D, Mouton PR. 2023. Novel stereological method for estimation of cell counts in 3D collagen scaffolds. Scientific Reports. 13(1), 7959. mla: Zavadakova, Anna, et al. “Novel Stereological Method for Estimation of Cell Counts in 3D Collagen Scaffolds.” Scientific Reports, vol. 13, no. 1, 7959, Springer Nature, 2023, doi:10.1038/s41598-023-35162-z. short: A. Zavadakova, L. Vistejnova, T. Belinova, F. Tichanek, D. Bilikova, P.R. Mouton, Scientific Reports 13 (2023). date_created: 2023-05-19T11:12:25Z date_published: 2023-05-17T00:00:00Z date_updated: 2023-08-01T14:46:06Z day: '17' ddc: - '570' department: - _id: Bio doi: 10.1038/s41598-023-35162-z external_id: isi: - '000995271600104' file: - access_level: open_access checksum: 8c1b769693ff4288df8376e59ad1176d content_type: application/pdf creator: dernst date_created: 2023-05-22T07:57:37Z date_updated: 2023-05-22T07:57:37Z file_id: '13047' file_name: 2023_ScientificReports_Zavadakova.pdf file_size: 3055077 relation: main_file success: 1 file_date_updated: 2023-05-22T07:57:37Z has_accepted_license: '1' intvolume: ' 13' isi: 1 issue: '1' keyword: - Multidisciplinary language: - iso: eng month: '05' oa: 1 oa_version: Published Version publication: Scientific Reports publication_identifier: issn: - 2045-2322 publication_status: published publisher: Springer Nature quality_controlled: '1' related_material: link: - relation: erratum url: https://doi.org/10.1038/s41598-023-37265-z scopus_import: '1' status: public title: Novel stereological method for estimation of cell counts in 3D collagen scaffolds tmp: image: /images/cc_by.png legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0) short: CC BY (4.0) type: journal_article user_id: 4359f0d1-fa6c-11eb-b949-802e58b17ae8 volume: 13 year: '2023' ...