Biology subjects

De Backer, J.

Publications and source records attributed to De Backer, J..

3 recordsLinked to original sources

bioRxiv2026

Integrated luminescence and phenotypic profiling for drug discovery in a zebrafish model of Marfan syndrome

BackgroundMarfan syndrome (MFS) is a life-threatening heritable connective tissue disorder caused by pathogenic variants in fibrillin-1, characterized by progressive cardiovascular disease. Current medical therapies slow disease progression but do not prevent major complications, underscoring the need for new treatment strategies and unbiased discovery approaches. MethodsWe used a zebrafish model of MFS lacking fibrillin-3 (fbn3-/-), which recapitulates key cardiovascular phenotypes including cardiac stress, valvular defects, arrhythmia, and aortic dilation. To enable sensitive, quantitative assessment of cardiac stress, we generated a novel transgenic zebrafish reporter expressing secreted nanoluciferase under control of the stress-responsive nppb promoter. This reporter was combined with morphological phenotyping and bulbus arteriosus (BA) imaging. We evaluated standard MFS therapies, targeted modulators of TGF-{beta} signaling, and performed an unbiased high-throughput drug screen of over 1 500 clinically approved compounds across multiple developmental treatment windows. Resultsfbn3-/- larvae exhibited markedly elevated nppb activity that correlated with phenotypic severity and peaked during stages of highest mortality. The nanoluciferase reporter provided a [~]1 000-fold dynamic range, substantially outperforming Firefly luciferase-based assays. Pharmacological inhibition of TGF-{beta} signaling produced transient or deleterious effects, while {beta}-blockers, losartan, and allopurinol failed to consistently improve cardiac stress, pericardial edema, or BA dilation. The unbiased high-throughput drug screen identified a small number of primary and secondary hits; however, none demonstrated reproducible phenotypic rescue upon rigorous multi-dose, multi-time window validation. ConclusionsThis study establishes a sensitive zebrafish-based platform for early, quantitative assessment of cardiovascular stress in MFS. Our findings highlight the limited efficacy of current therapies, the context-dependent nature of TGF-{beta} modulation, and the biological complexity underlying MFS pathogenesis. Although no definitive therapeutic candidates were identified, this work lays a robust foundation for expanded unbiased discovery efforts aimed at identifying disease-modifying interventions for MFS.

pharmacology and toxicology↗

bioRxiv2025

Systematic disruption of zebrafish fibrillin genes identifies a translational zebrafish model for Marfan syndrome

BackgroundFibrillins are essential components of the extracellular matrix. Marfan syndrome (MFS), the most common fibrillinopathy, is characterized by severe cardiovascular complications, including cardiac valve abnormalities, myocardial dysfunction, arrhythmias, and, most commonly, thoracic aortic disease. Unfortunately, no definitive medical cure is available. ObjectivesTo establish a zebrafish model of MFS, to enhance understanding of the cardiovascular consequences of fibrillin impairment and identify novel therapeutic targets. MethodsCRISPR/Cas9 technology was used to systematically target all zebrafish fibrillin genes. The cardiovascular phenotype was investigated using fluorescent microscopy at embryonic stages and cardiac ultrasound, histology, and synchrotron X-ray imaging in adults. RNA sequencing and drug testing were performed during early development. ResultsFibrillin-2b mutant (fbn2b-/-) zebrafish had a reproducible phenotype, with a subset of embryos showing endocardial detachment leading to early mortality. Interestingly, the remaining fbn2b-/- zebrafish developed dilation of the bulbus arteriosus, a structure analogous to the aortic root in humans, and survived normally to adulthood. Adult fbn2b-/- zebrafish displayed cardiac valve abnormalities. Transcriptomic analysis of fbn2b-/-embryos suggested the involvement of extracellular matrix remodeling and immune-related pathways. Administration of nebivolol and losartan did not improve the phenotype of fbn2b-/- larvae. Zebrafish lacking fibrillin-1 and/or fibrillin-2a did not show any phenotype. ConclusionOur fbn2b-/- zebrafish model recapitulates key aspects of human cardiovascular manifestations of MFS and can therefore be considered a novel relevant animal model for MFS. Studying this model allows us to broaden the knowledge of the underlying mechanisms of the disease and discover much-needed disease-specific treatment options. CONDENSED ABSTRACTFibrillin defects lead to severe cardiovascular complications in Marfan syndrome (MFS), including aortic dilation, dissection, and rupture. To model MFS, we generated zebrafish mutants lacking various fibrillin genes. Among these mutant lines, only fibrillin-2b-deficient zebrafish exhibited cardiovascular phenotypes mimicking human disease. Multimodal imaging revealed early cardiac defects, bulbus arteriosus dilation, and valve abnormalities. Transcriptomic analysis identified altered regulation of pathways related to extracellular matrix homeostasis and immune system activation. Compound testing demonstrated the models potential for drug discovery. This zebrafish model, recapitulating key cardiovascular features of MFS, provides a valuable platform to investigate disease mechanisms and identify novel treatment strategies.

genetics↗

bioRxiv2024

A novel genetic mouse model of fatal aortic dissection reveals massive inflammatory cell infiltration in the thoracic aorta

BackgroundAortic dissection (AD) is the separation of medial layers of the aorta and is a major cause of death in patients with connective tissue disorders such as Marfan syndrome. However, molecular triggers instigating AD, its temporospatial progression, and how vascular cells in each vessel layer interact and participate in the pathological process remain incompletely understood. To unravel the underlying molecular mechanism of AD, we generated a spontaneous AD mouse model. MethodsWe incorporated a novel missense variant (p.G234D) in FBN1, the gene for fibrillin-1, identified in a non-syndromic familial AD patient into mice using CRISPR/Cas9 system. We performed histopathological analyses of the aortic lesions by histology, immunofluorescence staining, electron microscopy, synchrotron-based imaging and single-cell (sc)RNA-sequencing. Biochemical analysis was performed to examine the binding capacity of mutant human FBN1G234D protein to latent Tgf{beta} binding proteins (LTBPs), and signaling pathways in the mutant aortic wall were examined by western blot analysis. Results50% of the Fbn1G234D/G234D mutant mice died within 5 weeks of age from multiple intimomedial tears that expanded longitudinally and progressed to aortic rupture accompanied by massive immune cell infiltration. scRNA-sequencing, validated by immunostaining, revealed a significant increase in MHC class II-positive pro-inflammatory macrophages and monocytes at the site of intima tears with upregulation of MMP2/9 and marked disruption of elastic lamina. Subendothelial matrices, such as type IV collagen and laminin, expanded into the medial layer, where fibronectin expression was highly upregulated. Fbn1G234D/G234D endothelial cells exhibited altered mechanosensing with loss of parallel alignment to blood flow and upregulation of VCAM-1 and ICAM-1, all of which likely contributed to the infiltration of immune cells. Biochemically, FBN1G234D lost the ability to bind to latent TGF{beta} binding protein (LTBP)-1, -2, and -4, resulting in the downregulation of TGF{beta} signaling in the aortic wall. ConclusionsWe show that dynamic interactions involving endothelial cells (ECs) and macrophages/monocytes in the intima, where the ECM microenvironment is altered with the reduced TGF{beta} signaling, contributes to the initiation of AD. Our novel AD mouse model provides a unique opportunity to identify target molecules involved in the intimomedial tears that can be utilized for development of therapeutic strategies.

pathology↗