Leonor Saúde

Instituto de Medicina Molecular

The vascular system is inefficiently repaired after spinal cord injury (SCI) in mammals, resulting in secondary tissue damage and immune deregulation that contribute to the limited functional recovery. Unlike mammals, zebrafish can repair the spinal cord (SC) and restore motility, but the vascular response to injury has not been investigated. Here,...

Traumatic spinal cord injury (SCI) initiates a cascade of cellular events, culminating in irreversible tissue loss and neuroinflammation. After the trauma, the blood vessels are destroyed. The blood-spinal cord barrier (BSCB), a physical barrier between the blood and spinal cord parenchyma, is disrupted, facilitating the infiltration of immune cell...

Cells with latent stem ability can contribute to mammalian tissue regeneration after damage. Whether the central nervous system (CNS) harbors such cells remains controversial. Here, we report that DNGR-1 lineage tracing in mice identifies an ependymal cell subset, wherein resides latent regenerative potential. We demonstrate that DNGR-1-lineage-tra...

The vascular system is inefficiently repaired after spinal cord injury in mammals, resulting in secondary tissue damage and immune deregulation that contribute to the limited functional recovery. Unlike mammals, zebrafish can repair the spinal cord and restore motility, but the vascular response to injury has not been investigated. Here we describe...

The cardiac developmental network has been associated with myocardial regenerative potential. However, the embryonic signals triggered following injury have yet to be fully elucidated. Nkx2.5 is a key causative transcription factor associated with human congenital heart disease and one of the earliest markers of cardiac progenitors, thus it serves...

Cellular senescence is a highly complex and programmed cellular state with diverse and, at times, conflicting physiological and pathological roles across the lifespan of an organism. Initially considered a cell culture artifact, senescence evolved from an age-related circumstance to an intricate cellular defense mechanism in response to stress, imp...

Persistent senescent cells (SCs) are known to underlie aging-related chronic disorders, but it is now recognized that SCs may be at the center of tissue remodeling events, namely during development or organ repair. In this study, we show that two distinct senescence profiles are induced in the context of a spinal cord injury between the regenerativ...

The vascular bioactivity/safety of nanomaterials is typically evaluated by animal testing, which is of low throughput and does not account for biological differences between animals and humans such as ageing, metabolism and disease profiles. The development of personalized human in vitro platforms to evaluate the interaction of nanomaterials with t...

Persistent senescent cells (SCs) are known to underlie ageing-related chronic disorders, but it is now recognized that SCs may be at the center of tissue remodeling events, namely during development or organ repair. Here we show that two distinct senescence profiles are induced in the context of a spinal cord injury between the regenerating zebrafi...

Low doses of ionizing radiation (LDIR) activate endothelial cells inducing angiogenesis. In zebrafish, LDIR induce vessel formation in the sub-intestinal vessels during post-embryonic development and enhance the inter-ray vessel density in adult fin regeneration. Since angiogenesis is a crucial process involved in both post-embryonic development an...

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Spinal cord injury (SCI) is a complex condition, with limited therapeutic options, that results in sensory and motor disabilities. To boost discovery of novel therapeutics, we designed a simple and efficient drug screening platform. This innovative approach allows to determine locomotor rescue properties of small molecules in a zebrafish (Danio rer...

The formation of distinct 3'UTRs through alternative polyadenylation is a mechanism of gene expression regulation that has been implicated in many physiological and pathological processes. However, its functions in the context of vertebrate embryonic development have been largely unaddressed, in particular with a gene-specific focus. Here we show t...

Background: Dmrt2a is a zinc finger like transcription factor with several roles during zebrafish early development: left-right asymmetry, synchronisation of the somite clock genes and fast muscle differentiation. Despite the described functions, Dmrt2a mechanism of action is unknown. Therefore, with this work, we propose to identify Dmrt2a downst...

Zebrafish are able to regenerate the spinal cord and recover motor and sensory functions upon severe injury, through the activation of cells located at the ependymal canal. Here, we show that cells surrounding the ependymal canal in the adult zebrafish spinal cord express Foxj1a. We demonstrate that ependymal cells express Foxj1a from their birth i...

GO annotation for gene groups obtained with the R package clusterProfiler.

Contains data on foxj1a gene expression by in situ hybridization and by qPCR.

Contains data about the number of motile and immotile cilia from single embryos along development from 3 ss to 8 ss (n = 4).

Contains data on the relative expression levels of several genes by quantitative PCR. It shows comparisons between dld−/− mutants and wild type embryos.

Contains data from the native particles tracked to generate the flow maps on Figure 2. Shows the evaluation of the motile and immotile cilia by transmitted light microscopy with a high speed video-camera and the corresponding cilia beat frequency for the motile cilia.

Contains data on her12 positive DFC number and its anterior posterior location within the DFC cluster.

Microarray data. Excel file that contains Table S1a - List of 706 genes with significantly altered transcription. This list contains 706 genes with a fold change in transcription higher than 2, in the DFCs from dld−/− mutant zebrafish embryos. Table S1b – List of motility associated genes from the Table S1a that have been associated with cilia in t...

Contains the R script for creating and analysing the flow maps.

Provides data on the coordinates of immotile cilia denoting posterior to anterior transitions.

Relative expression levels of dnah7, her12, dnah9, rfx4 and foxj1a by qPCR for different Notch signalling manipulations. Contains data and statistical tests for the number of motile cilia per KV.

Provides data on flow speed and CBF upon her12 overexpression. Shows data for dand5 pattern and organ situs scoring per embryo. Shows the data for cilia length measurements in 3D and the evaluation of motile and immotile cilia localization according to the anterior – posterior axis of each stack of images.

Foxj1a is necessary and sufficient to specify motile cilia. Using transcriptional studies and slow-scan two-photon live imaging capable of identifying the number of motile and immotile cilia, we now established that the final number of motile cilia depends on Notch signalling (NS). We found that despite all left-right organizer (LRO) cells express...

The use of gold nanoparticles for effective gene silencing has demonstrated its potential as a tool for gene expression experiments and for the treatment of several diseases. Here, we used a gold nanobeacon designed to specifically silence the enhanced green fluorescence protein (EGFP) mRNA in embryos of a fli-EGFP transgenic zebrafish line, while...

The European Zebrafish Principal Investigator Meeting (EZPM) is an ideal forum for group leaders using this fantastic animal model not only to discuss science but also to strengthen their interactions, to push forward technological advances, and to define guidelines for the use of this fish in research. The city of Lisbon (Portugal) was voted by th...

The stereotypic left-right (LR) asymmetric distribution of internal organs is due to an asymmetric molecular cascade in the lateral plate mesoderm (LPM) that is originated at the embryonic node. In chicken embryos, molecular asymmetries at Hensen's node are created by leftward cell movements that occur transiently. What terminates these movements,...

Accurate regulation of Notch signalling is central for developmental processes in a variety of tissues, but its function in pectoral fin development in zebrafish is still unknown. Here we show that core elements necessary for a functional Notch pathway are expressed in developing pectoral fins in or near prospective muscle territories. Blocking Not...

Expression pattern of Notch signalling pathway genes at early and late time points of pectoral fin development. Expression of jagged2 (n = 25) (A), notch1a (n = 27) (B), notch2 (n = 20) (C), notch3 (n = 25) (D), her6 (n = 22) (E), her7 (n = 20) (F) and her13.2 (n = 20) (G) can be detected in the entire pectoral fin at 36 hpf. Later in development a...

Proximal-distal and anterior-posterior patterning and Hox identity are not affected in mibta52b mutants. The Fgf signalling components fgf8 (n = 15) (A, A’) and fgf24 (n = 12) (C, C’) are expressed in the apical ectodermal fold of 48 hpf sibling embryos. The same pattern of expression is observed in the pectoral fins of mibta52b mutants (n = 12) (B...

Zebrafish (Danio rerio) has a remarkable capacity to regenerate many organs and tissues. During larval stages the fin fold allows the possibility of performing long time-lapse imaging making this system very appealing to study the relationships between tissue movements, cell migration and proliferation necessary for the regeneration process. Throug...

Expression pattern of the 2 dpf transgenic osteopontin:eGFP. At this stage of development, this transgenic has labeled the fin fold mesenchymal cells, but also other mesenchymal cells that are spread out along the midline and somites. Besides these, the pectoral fin, the eye and the brain are also GFP positive. (TIF)

Wound healing process of 2 dpf Alpha-Catenin-Citrine transgenic fish. 1 hour movie of a 2 dpf 5 minpa larva. Anterior is to the left. Scale bar corresponds to 50 µm. (MOV)

Alpha-Catenin is present in the actin cable during wound healing. 3D reconstruction of a representative immunostaining against anti-GFP antibody to detect alpha-catenin (green) and phalloidin to detect actin (red) in 2 dpf 5 minpa alpha-catenin-Citrine (ctnna-Citrine) transgenic larva. n = 5 larvae. (MOV)

2 dpf Uncut control double transgenic EF1α:mKO2-zCdt1;osteopontin:eGFP. 6 hours movie of an uncut 2 dpf larva. Scale bar corresponds to 50 µm. (MOV)

Wound healing process of 2 dpf actb1:myl12.1-eGFP transgenic fish. 3 hours movie of a 2 dpf 5 minpa larva. Anterior is to the left. Scale bar corresponds to 50 µm. (MOV)

Wound healing process of 2 dpf Utrophin-GFP injected fish. 1 hour movie of a 2 dpf 5 minpa larva. Anterior is to the left. Scale bar corresponds to 50 µm. (MOV)

Apoptosis is not present during fin fold regeneration. Representative immunofluorescence with anti-active Caspase3 antibody in uncut and amputated larvae of 3 dpf (A–B), 4 dpf (C–D) and 5 dpf (E–F). Arrow indicates the presence of an apoptotic cell. n = 5 larvae per condition. Scale bar corresponds to 50 µm in all images. (TIF)

Detailed distribution of cell division angles in the 4 regions of the fin fold. (TIF)

The mesenchymal cells are maintained in G0-G1 phases of the cell cycle regardless of an amputation. Live imaging representative images of double transgenic EF1α:mKO2-zCdt1;osteopontin:eGFP larvae during several stages of the regenerative process and their respective controls. A,C,E are uncut (3 dpf, 4 dpf and 5 dpf respectively) and age matched con...

The fin fold regeneration dynamics are independent of the size of amputation. Representative brightfield live images of AB (A–L) and TU (M–X) larvae during several stages of the regenerative process and their respective age-matched uncut controls. Larvae were subjected to different amputation planes (Regular, Half-size and Diagonal cuts) and follow...

2 dpf 30 minpa double transgenic EF1α:mKO2-zCdt1;osteopontin:eGFP. 6 hours movie of a 2 dpf larva 30 minpa. Scale bar corresponds to 50 µm. (MOV)

The mesenchymal cells are polarized. A Representative immunostaining with anti-GFP and anti-γTubulin antibodies in 2 dpf uncut transgenic osteopontin:eGFP larvae (single frame). B–C Representative immunostaining with anti-GFP anti-γTubulin antibodies in amputated transgenic osteopontin:eGFP larvae of 2 dpf 1 hpa and 3 dpf 1 dpa (single frames). A'–...

Somites are formed from the presomitic mesoderm (PSM) and give rise to the axial skeleton and skeletal muscles. The PSM is dynamic; somites are generated at the anterior end, while the posterior end is continually renewed with new cells entering from the tailbud progenitor region. Which genes control the conversion of tailbud progenitors into PSM a...

Background Zebrafish has emerged as a powerful model organism to study the process of regeneration. This teleost fish has the ability to regenerate various tissues and organs like the heart, spinal cord, retina and fins. In this study, we took advantage of the existence of an excellent morphological reference in the zebrafish caudal fin, the bony r...

Fgf signalling does not seem to play a role in the determination of the proximal-distal position of the bifurcation. Transgenic hsp70:dn-fgfr1 fish were amputated 1 segment proximal to the bifurcation and heat-shocked at: 35°C for 1 hour, every other day, from day 2 post amputation until day 8 post amputation; 36°C for 1 hour daily, during 3 days,...

Mesp proteins play crucial roles in the formation of heart, vasculature and somites during vertebrate embryogenesis. We have used phylogenetic and genomic analysis, combined with qRT-PCR and in situ hybridization, to characterize two novel additional mesp genes in zebrafish, mesp-ab and mesp-bb, and describe their expression pattern in wild type an...

The zebrafish has the capacity to regenerate many tissues and organs. The caudal fin is one of the most convenient tissues to approach experimentally due to its accessibility, simple structure and fast regeneration. In this work we investigate how the regenerative capacity is affected by recurrent fin amputations and by experimental manipulations t...

Members of the Dmrt family, generally associated with sex determination, were shown to be involved in several other functions during embryonic development. Dmrt2 has been studied in the context of zebrafish development where, due to a duplication event, two paralog genes dmrt2a and dmrt2b are present. Both zebrafish dmrt2a/terra and dmrt2b are impo...

The importance of cilia in embryonic development and adult physiology is emphasized by human ciliopathies. Despite its relevance, molecular signalling pathways behind cilia formation are poorly understood. We show that Notch signalling is a key pathway for cilia length control. In deltaD zebrafish mutants, cilia length is reduced in Kupffer's vesic...

The formation of a perfect vertebrate body plan poses many questions that thrill developmental biologists. Special attention has been given to the symmetric segmental patterning that allows the formation of the vertebrae and skeletal muscles. These segmented structures derive from bilaterally symmetric units called somites, which are formed under t...

At the European Council meeting in Lisbon in 2000, the European Union (EU) established the strategic goal of becoming “the most dynamic and competitive knowledge‐based economy in the world” by 2010 (Fontaine, 2000). To achieve this, each EU member state would be required to increase its investment in research and development (R&D) to 3% of its Gros...

The 1st Meeting of the Portuguese Society for Developmental Biology (SPBD ­ Sociedade Portuguesa de Biologia do Desenvolvimento) (Fig. 1) was held during two sunny autumn days at the Instituto Gulbenkian de Ciencia (IGC) located in the beautiful seaside city of Oeiras. This small meeting provided unprecedented conditions for Portuguese Developmenta...

To establish the vertebrate body plan, it is fundamental to create left-right asymmetry in the lateral-plate mesoderm to correctly position the organs. However, it is also crucial to maintain symmetry between the left and the right sides of the presomitic mesoderm, ensuring the allocation of symmetrical body structures, such as the axial skeleton a...