| 1. |
- Almstedt, Elin, et al.
(author)
-
Real-time evaluation of glioblastoma growth in patient-specific zebrafish xenografts
- 2021
-
In: Neuro-Oncology. - : Oxford University Press. - 1522-8517 .- 1523-5866. ; 24:5, s. 726-738
-
Journal article (peer-reviewed)abstract
- Background: Patient-derived xenograft (PDX) models of glioblastoma (GBM) are a central tool for neuro-oncology research and drug development, enabling the detection of patient-specific differences in growth, and in vivo drug response. However, existing PDX models are not well suited for large-scale or automated studies. Thus, here, we investigate if a fast zebrafish-based PDX model, supported by longitudinal, AI-driven image analysis, can recapitulate key aspects of glioblastoma growth and enable case-comparative drug testing.Methods: We engrafted 11 GFP-tagged patient-derived GBM IDH wild-type cell cultures (PDCs) into 1-day-old zebrafish embryos, and monitored fish with 96-well live microscopy and convolutional neural network analysis. Using light-sheet imaging of whole embryos, we analyzed further the invasive growth of tumor cells.Results: Our pipeline enables automatic and robust longitudinal observation of tumor growth and survival of individual fish. The 11 PDCs expressed growth, invasion and survival heterogeneity, and tumor initiation correlated strongly with matched mouse PDX counterparts (Spearman R = 0.89, p < 0.001). Three PDCs showed a high degree of association between grafted tumor cells and host blood vessels, suggesting a perivascular invasion phenotype. In vivo evaluation of the drug marizomib, currently in clinical trials for GBM, showed an effect on fish survival corresponding to PDC in vitro and in vivo marizomib sensitivity.Conclusions: Zebrafish xenografts of GBM, monitored by AI methods in an automated process, present a scalable alternative to mouse xenograft models for the study of glioblastoma tumor initiation, growth, and invasion, applicable to patient-specific drug evaluation.
|
|
| 2. |
- Arnold, Hannah, et al.
(author)
-
mafba and mafbb differentially regulate lymphatic endothelial cell migration in topographically distinct manners
- 2022
-
In: Cell Reports. - : Elsevier. - 2211-1247. ; 39:12
-
Journal article (peer-reviewed)abstract
- Lymphangiogenesis, formation of lymphatic vessels from pre-existing vessels, is a dynamic process that requires cell migration. Regardless of location, migrating lymphatic endothelial cell (LEC) progenitors probe their surroundings to form the lymphatic network. Lymphatic-development regulation requires the transcription factor MAFB in different species. Zebrafish Mafba, expressed in LEC progenitors, is essential for their migration in the trunk. However, the transcriptional mechanism that orchestrates LEC migration in different lymphatic endothelial beds remains elusive. Here, we uncover topographically different requirements of the two paralogs, Mafba and Mafbb, for LEC migration. Both mafba and mafbb are necessary for facial lymphatic development, but mafbb is dispensable for trunk lymphatic development. On the molecular level, we demonstrate a regulatory network where Vegfc-Vegfd-SoxF-Mafba-Mafbb is essential in facial lymphangiogenesis. We identify that mafba and mafbb tune the directionality of LEC migration and vessel morphogenesis that is ultimately necessary for lymphatic function.
|
|
| 3. |
- Arnold, Hannah, et al.
(author)
-
Mafba and Mafbb Differentially Regulate Lymphatic Endothelial Cell Migration in Topographically Distinct Manners
- 2021
-
In: SSRN Electronic Journal. - : Elsevier. - 1556-5068.
-
Journal article (peer-reviewed)abstract
- Lymphangiogenesis is the formation of lymphatic vessels from pre-existing vessels, a dynamic process that requires cell migration. Regardless of location, lymphatic endothelial cell (LEC) progenitors probe their surroundings while migrating to form the lymphatic network. Lymphatic development regulation depends on the transcription factor MAFB in different species. Zebrafish Mafba, expressed in LEC progenitors, is essential for their migration in the trunk. However, the transcriptional mechanism that orchestrate LEC migration in different lymphatic endothelial beds remains elusive. Here, we uncover topographically different requirements of the two paralogues, Mafba and Mafbb, for lymphatic cell migration. Both mafba and mafbb are necessary for facial lymphatic development, but mafbb is dispensable for trunk lymphatic development. On the molecular level, we demonstrate a regulatory network where Vegfc-Vegfd-SoxF-Mafba-Mafbb are essential in the facial lymphangiogenesis. We identify that mafba and mafbb fine-tune the directionality of LEC migration and vessel morphogenesis that is ultimately necessary for lymphatic function.
|
|
| 4. |
- Gloger, Marleen, et al.
(author)
-
Spatio-temporal control of lymphatic endothelial cell proliferation during embryonic development
-
Other publication (other academic/artistic)abstract
- Cell proliferation is central to proper formation of functional and healthy organs. During embryogenesis, lymphatic vessels are derived from progenitor pools that undergo cell proliferation to ensure the emergence of the accurate lymphatic cell number. Altered lymphatic endothelial cell (LEC) proliferation can lead to vessel hypo- or hyperplasia, which is the underlying basis of lymphoedema. Currently, there is no cure for these diseases. Although Vegfc-Vegfr3 (Flt4) signalling has been uncovered as the apical mitogenic pathway in LEC proliferation, there is a knowledge-gap of whether this pathway induces the basic cell cycle machinery or if other lymphatic lineage-specific components exist to regulate cell proliferation. Current studies showed, that in ddx21 mutants specifically LEC emergence and proliferation upon Vegfc-Vegfr3 signalling appears to be disrupted; whereas venous endothelial cell (VEC) proliferation and venous intersegmental vessel formation was unaffected. This suggests that there might be a lineage-specific molecular code that ensures tightly regulated LEC proliferation. To determine how proliferation contributes to lymphatic development, we have performed a time-course analysis and quantification of the total number of venous endothelial cells (VECs) and LECs in the whole zebrafish. This revealed three crucial time points where VEC and LEC cell number expansion takes place during larval stages of developmental. Detailed analysis for VECs and LECs confirmed that this was specific to all lymphatic beds developing at these time points. Using 5-ethynyl-2-deoxyuridine (EdU), we confirmed that the expansion of the LEC number is due to active proliferation. Analysis of the VEC and LEC single-cell RNA (scRNA) sequencing data identified novel molecular factors regulating LEC proliferation. Suggesting that this LEC lineage-specific proliferation gene signature acts in conjunction with the basic cell cycle machinery and regulates lymphatic vessel development in a spatio-temporal controlled manner. We believe that this study provides knowledge that will be highly relevant for understanding the basis of lymphoedema.
|
|
| 5. |
- Koltowska, Katarzyna, et al.
(author)
-
The RNA helicase Ddx21 controls Vegfc-driven developmental lymphangiogenesis by balancing endothelial cell ribosome biogenesis and p53 function
- 2021
-
In: Nature Cell Biology. - : Springer Nature. - 1465-7392 .- 1476-4679. ; 23:11, s. 1136-1147
-
Journal article (peer-reviewed)abstract
- Hogan and colleagues report that the RNA helicase Ddx21 mediates Vegfc-stimulated lymphangiogenesis during zebrafish development through controlling rDNA transcription and ribosome biogenesis in endothelial cells. The development of a functional vasculature requires the coordinated control of cell fate, lineage differentiation and network growth. Cellular proliferation is spatiotemporally regulated in developing vessels, but how this is orchestrated in different lineages is unknown. Here, using a zebrafish genetic screen for lymphatic-deficient mutants, we uncover a mutant for the RNA helicase Ddx21. Ddx21 cell-autonomously regulates lymphatic vessel development. An established regulator of ribosomal RNA synthesis and ribosome biogenesis, Ddx21 is enriched in sprouting venous endothelial cells in response to Vegfc-Flt4 signalling. Ddx21 function is essential for Vegfc-Flt4-driven endothelial cell proliferation. In the absence of Ddx21, endothelial cells show reduced ribosome biogenesis, p53 and p21 upregulation and cell cycle arrest that blocks lymphangiogenesis. Thus, Ddx21 coordinates the lymphatic endothelial cell response to Vegfc-Flt4 signalling by balancing ribosome biogenesis and p53 function. This mechanism may be targetable in diseases of excessive lymphangiogenesis such as cancer metastasis or lymphatic malformation.
|
|
| 6. |
|
|
| 7. |
- Peng, Di, et al.
(author)
-
Proper migration of lymphatic endothelial cells requires survival and guidance cues from arterial mural cells
- 2022
-
In: eLIFE. - : eLife Sciences Publications Ltd. - 2050-084X. ; 11
-
Journal article (peer-reviewed)abstract
- The migration of lymphatic endothelial cells (LECs) is key for the development of the complex and vast lymphatic vascular network that pervades most tissues in an organism. In zebrafish, arterial intersegmental vessels together with chemokines have been shown to promote lymphatic cell migration from the horizontal myoseptum (HM). We observed that emergence of mural cells around the intersegmental arteries coincides with lymphatic departure from HM which raised the possibility that arterial mural cells promote LEC migration. Our live imaging and cell ablation experiments revealed that LECs migrate slower and fail to establish the lymphatic vascular network in the absence of arterial mural cells. We determined that mural cells are a source for the C-X-C motif chemokine 12 (Cxcl12a and Cxcl12b), vascular endothelial growth factor C (Vegfc) and collagen and calcium-binding EGF domain-containing protein 1 (Ccbe1). We showed that chemokine and growth factor signalling function cooperatively to induce robust LEC migration. Specifically, Vegfc-Vegfr3 signalling, but not chemokines, induces extracellular signal-regulated kinase (ERK) activation in LECs, and has an additional pro-survival role in LECs during the migration. Together, the identification of mural cells as a source for signals that guide LEC migration and survival will be important in the future design for rebuilding lymphatic vessels in disease contexts.
|
|
| 8. |
- Richards, Mark, et al.
(author)
-
Claudin5 protects the peripheral endothelial barrier in an organ and vessel-type-specific manner
- 2022
-
In: eLIFE. - : eLife Sciences Publications Ltd. - 2050-084X. ; 11
-
Journal article (peer-reviewed)abstract
- Dysfunctional and leaky blood vessels resulting from disruption of the endothelial cell (EC) barrier accompanies numerous diseases. The EC barrier is established through endothelial cell tight and adherens junctions. However, the expression pattern and precise contribution of different junctional proteins to the EC barrier is poorly understood. Here, we focus on organs with continuous endothelium to identify structural and functional in vivo characteristics of the EC barrier. Assembly of multiple single-cell RNAseq datasets into a single integrated database revealed the variability and commonalities of EC barrier patterning. Across tissues, Claudin5 exhibited diminishing expression along the arteriovenous axis, correlating with EC barrier integrity. Functional analysis identified tissue-specific differences in leakage properties and response to the leakage agonist histamine. Loss of Claudin5 enhanced histamine-induced leakage in an organotypic and vessel type-specific manner in an inducible, EC-specific, knock-out mouse. Mechanistically, Claudin5 loss left junction ultrastructure unaffected but altered its composition, with concomitant loss of zonula occludens-1 and upregulation of VE-Cadherin expression. These findings uncover the organ-specific organisation of the EC barrier and distinct importance of Claudin5 in different vascular beds, providing insights to modify EC barrier stability in a targeted, organ-specific manner.
|
|
| 9. |
- Vallecillo-Garcia, Pedro, et al.
(author)
-
A local subset of mesenchymal cells expressing the transcription factor Osr1 orchestrates lymph node initiation
- 2023
-
In: Immunity. - : Cell Press. - 1074-7613 .- 1097-4180. ; 56:6, s. 1204-
-
Journal article (peer-reviewed)abstract
- During development, lymph node (LN) initiation is coordinated by lymphoid tissue organizer (LTo) cells that attract lymphoid tissue inducer (LTi) cells at strategic positions within the embryo. The identity and function of LTo cells during the initial attraction of LTi cells remain poorly understood. Using lineage tracing, we demon-strated that a subset of Osr1-expressing cells was mesenchymal LTo progenitors. By investigating the het-erogeneity of Osr1+ cells, we uncovered distinct mesenchymal LTo signatures at diverse anatomical loca-tions, identifying a common progenitor of mesenchymal LTos and LN-associated adipose tissue. Osr1 was essential for LN initiation, driving the commitment of mesenchymal LTo cells independent of neural ret-inoic acid, and for LN-associated lymphatic vasculature assembly. The combined action of chemokines CXCL13 and CCL21 was required for LN initiation. Our results redefine the role and identity of mesenchymal organizer cells and unify current views by proposing a model of cooperative cell function in LN initiation.
|
|
