Biology subjects

Eley, L.

Publications and source records attributed to Eley, L..

3 recordsLinked to original sources

bioRxiv2025

Remodelling of supernumerary leaflet primordia leads to bicuspid aortic valve (BAV) caused by loss of primary cilia

AimsBicuspid aortic valve (BAV), where two valve leaflets are found instead of the usual three, affects 1-2% of the general population and is associated with significant morbidity and mortality. Despite its frequency, the majority of cases remain unexplained. This is, at least in part, because there are two types of valve leaflet primordia: endocardial cushions and intercalated valve swellings (ICVS). Moreover, multiple progenitors make distinct contribution to the formation of these primordia. Genomic studies in mouse and human have suggested a correlation between BAV and malfunctional primary cilia. However, the precise requirement for cilia during early embryonic valvulogenesis remains unknown. Methods and resultsHere, we disrupted primary cilia by deleting the ciliary gene Ift88 in the main progenitor cells forming the aortic valve using specific Cre drivers: Wnt1-Cre for neural crest cells, Isl1-Cre for second heart field cells (SHF); Tie2-Cre for endocardial-derived cells and Tnnt2-Cre for direct-differentiating SHF in the ICVS. Loss of Ift88, and thus primary cilia, from neural crest cells and endocardium did not impact aortic valve formation. However, primary cilia are essential in SHF cells for aortic valve leaflet formation, with over half of Ift88f/f;Isl1-Cre mutants presenting with BAV. As the valve leaflets are forming, 50% of the Ift88f/f;Isl1-Cre mutants have two small leaflets in the position of the usual posterior leaflet, meaning that at this stage the aortic valve is quadricuspid, which then remodels to BAV by E15.5. Mechanistic studies demonstrate premature differentiation of SHF cells as the ICVS form, leading to the formation of a broadened ICVS that forms two posterior leaflet precursors. This abnormality in the formation of the ICVS is associated with disruption of Notch-Jag1 signalling pathway, with Jag1f/f;Isl1-Cre mutants presenting with a similar phenotype. ConclusionsThese data show that primary cilia, via the Notch-Jag1 signalling pathway, regulate differentiation of SHF cells in the aortic valve primordia. Additionally, we identify a mechanistic link between the developmental basis of quadricuspid and bicuspid arterial valve leaflets. Translational PerspectiveSeveral genomic studies in human and mouse have suggested that disruption of cilia-related genes may be a significant cause of CHD. Although there is limited data from animal models to suggest a link between cilia and bicuspid aortic valve (BAV), the mechanisms underpinning BAV formation during early valvulogenesis have not been described. Here, we established a potential mechanism underpinning BAV formation, highlighting a role for primary cilia in a subset of valve interstitial cells (VIC) derived from second heart field progenitors. Loss of cilia altered VIC differentiation and valvulogenesis. This study confirms that disruption of cilial formation and/or function can lead to arterial valve defects and could pave the way to finding therapies for patient benefit.

developmental biology↗

bioRxiv2024

Zebrafish arterial valve development occurs through direct differentiation of second heart field progenitors

AimsBicuspid Aortic Valve (BAV) is the most common congenital heart defect, affecting at least 2% of the population. The embryonic origins of BAV remain poorly understood, limiting the identification of assays for validating patient variants and ultimately causative genes for BAV. In both human and mouse, the left and right leaflets of the arterial valves arise from the outflow tract cushions, with interstitial cells originating from neural crest cells and endocardial-to-mesenchymal transition (EndoMT). In contrast, an EndoMT-independent mechanism of direct differentiation by cardiac progenitors from the second heart field (SHF) is responsible for the formation of the anterior and posterior leaflets. Defects in either of these developmental mechanisms can result in BAV. Although zebrafish have been suggested as a model for human variant testing, their naturally bicuspid arterial valve has not been considered suitable for understanding human arterial valve development. Here, we have set out to investigate to what extent the processes involved in arterial valve development are conserved in zebrafish and ultimately, whether functional testing of BAV variants could be carried out in zebrafish. Methods and ResultsUsing a combination of live imaging, immunohistochemistry and Cre-mediated lineage tracing, we show that the zebrafish arterial valve primordia develop directly from undifferentiated SHF progenitors with no contribution from EndoMT or neural crest, in keeping with the human and mouse anterior and posterior leaflets. Moreover, once formed, these primordia share common subsequent developmental events with all three mammalian arterial valve leaflets. ConclusionsOur work highlights a conserved ancestral mechanism of arterial leaflet formation from the SHF and identifies that development of the zebrafish arterial valve is distinct from that of the atrioventricular valve. Crucially, this confirms the utility of zebrafish for understanding the development of specific BAV subtypes and arterial valve dysplasia, offering potential for high-throughput variant testing. Translational PerspectiveLarge genomic studies of patients with Bicuspid Aortic Valve (BAV) have identified numerous variants predicted to be causative, yet due to a lack of suitable, in vivo functional assays, advancement of genetic testing, discussion of risk to family members and accurate prognosis is not yet widely possible. Here, we show that zebrafish demonstrate a high level of conservation in arterial valve development with the intercalated leaflets in human, establishing zebrafish as a suitable in vivo model that can begin to overcome the disconnect between clinical genetics and developmental biology.

developmental biology↗

bioRxiv2023

Spatial transcriptomics reveals novel genes during the remodelling of the embryonic human arterial valves

Abnormalities of the arterial valves, including bicuspid aortic valve (BAV) are amongst the most common congenital defects and are a significant cause of morbidity as well as predisposition to disease in later life. Despite this, and compounded by their small size and relative inaccessibility, there is still much to understand about how the arterial valves form and remodel during embryogenesis, both at the morphological and genetic level. Here we set out to address this in human embryos, using Spatial Transcriptomics (ST). We show that ST can be used to investigate the transcriptome of the developing arterial valves, circumventing the problems of accurately dissecting out these tiny structures from the developing embryo. We show that the transcriptome of CS16 and CS19 arterial valves overlap considerably, despite being several days apart in terms of human gestation, and that expression data confirm that the great majority of the most differentially expressed genes are valve-specific. Moreover, we show that the transcriptome of the human arterial valves overlaps with that of mouse atrioventricular valves from a range of gestations, validating our dataset but also highlighting novel genes, including four that are not found in the mouse genome and have not previously been linked to valve development. Importantly, our data suggests that valve transcriptomes are under-represented when using commonly used databases to filter for genes important in cardiac development; this means that causative variants in valve-related genes may be excluded during filtering for genomic data analyses for, for example, BAV. Finally, we highlight "novel" pathways that likely play important roles in arterial valve development, showing that mouse knockouts of RBP1 have arterial valve defects. Thus, this study has confirmed the utility of ST for studies of the developing heart valves and broadens our knowledge of the genes and signalling pathways important in human valve development. Non-Technical SummaryCongenital heart defects, particularly those affecting the valves and septa of the heart, are very common. Despite this, few gene variants have been confirmed as disease-causing in human congenital heart (including valve) disease patients. Here we utilise spatial transcriptomics technology, which allows the identification of genes expressed in tissue slices, on embryonic human heart valves and identify a gene dataset that is human arterial valve-specific. We confirm the localisation of key novel genes to the arterial valves and highlight the relevance of the dataset by showing that mice mutant for RBP1, a novel gene identified as being highly differentially expressed in our valve dataset, have previously unidentified arterial valve defects. Using commonly used bioinformatic databases we show that filtering patient genomic data using these terms would likely exclude valve genes and thus may not identify the causative genes. Thus, we confirm that spatial transcriptomics technology can be used to study gene expression in tiny structures such as the developing heart valves and provide a new human embryonic valve dataset that can be used in future genomic studies of patients with congenital valve defects.

developmental biology↗