Thoracaal aorta aneurysma

Investigating thoracic aortic aneurysm pathogenesis at single-cell resolution.

Thoracic aortic aneurysm (TAA) is an abnormal widening of the aorta in the chest, caused by the weakening of the aortic wall. TAAs can lead to rupture or dissection, a devastating complication with a mortality rate of 50%. Despite considerable efforts to gain insights on the molecular mechanisms underlying TAAs, there is currently no therapy that effectively stops or reverses TAA development. Single-cell RNA sequencing (scRNA-seq) is emerging as a ground-breaking technology to investigate gene expression at single-cell level and is opening new avenues to discover yet unexplored disease pathways. In my project, I will apply this technique to investigate a novel TAA disorder caused by biallelic pathogenic variants in the IPO8 gene, recently discovered in our Cardiogenomics research group. I will search for differentially expressed genes (DEGs) within the different aortic cell populations from an Ipo8^-/- mouse model that recapitulates the human aortic aneurysmal phenotype. I will also investigate shared DEGs between Ipo8^-/- mice and additional TAAs mouse models to find convergent disease pathways in clinically related TAA disorders. Subsequently, I will validate the role of the identified candidate culprits in mouse TAA development in a human setting, by using CRISPR-inhibition or -activation in iPSCs derived vascular smooth muscle cells or endothelial cells. The predicted outcomes will potentially pinpoint novel TAA drivers and hence, unveil potential new therapeutic targets.

PhD student: Irene Valdivia Callejon
Promotors: Aline Verstraeten, Bart Loeys & Josephina Meester

Converging mechanisms and biomarkers for thoracic aortic aneurysm and dissection.

Thoracic aortic aneurysms and dissections are a devastating cause of cardiovascular death at young age. Since only a fraction of the underlying pathogenetic factors have been elucidated and current treatments cannot halt the disease process, our first challenge is to dissect the convergent transcriptomic landscape of the aneurysmal aorta by using bulk RNA sequencing. This robust approach will unequivocally provide valuable insights into converging disease pathways that enable development of causal therapies.

Rapid, uncostly and large scale screening is paramount for early detection of aortic aneurysm, which is asymptomatic until dissection or rupture occurs. Fortunately, the transcriptomic landscape of the diseased aorta will also allow targeted development of blood biomarkers, seeing that expression patterns of diseased tissue leave an imprint in circulation. Multicenter blood sample collection from TAA patients is ongoing for this purpose, and mRNA, microRNA and protein analyses will be performed on these samples in the near future.

PhD student: Jotte Rodrigues Bento
Promotors: Bart Loeys, Aline Verstraeten & Josephina Meester

Discovery of genetic modifiers of the phenotypical cardiovascular variability in Marfan syndrome to pave the road to individualized treatment. protocols

Marfan syndrome (MFS) is an autosomal dominant connective tissue disorder with pleiotropic ocular, skeletal and cardiovascular manifestations. Morbidity and mortality are mostly determined by aortic root aneurysm, dissection and rupture. Although mutations in FBN1, coding for fibrillin-1, are the sole genetic MFS cause, there is a poor correlation between the MFS phenotype and the nature or location of the FBN1 variant. Wide intra- and interfamilial phenotypic variability, ranging from completely asymptomatic to sudden death at young age, is observed. The precise mechanisms underlying this variability remain elusive. In this project, I have selected an innovative strategy to fully understand the functional effects of the FBN1 mutation and discover genetic modifiers of MFS aortopathy with the following objectives: (1) CRISPR/Cas9 correction of the recurrent FBN1 p.Ile2585Thr in patient-derived iPSC-VSMCs and functional comparison to FBN1 mutation and control iPSC-VSMCs. (2) Whole genome sequencing, and RNA-seq of patient iPSCVSMCs at the extreme ends of the phenotypical spectrum for genetic modifier identification. (3) CRISPR-modification for validation of their modifying capacities. The understanding of the functional effects of the FBN1 mutation and the identification of genetic modifiers will advance the knowledge on aortopathy-mechanisms beyond current understanding, it will allow to individualize treatment protocols and will offer new leads to novel therapeutic targets.

PhD student: Lotte Van Den Heuvel
Promotors: Bart Loeys, Aline Verstraeten & Josephina Meester

In search of genetic modifiers for aortopathy in Loeys-Dietz syndrome families with a SMAD3 mutation.

Thoracic aortic aneurysm (TAA) results from progressive dilatation of the aorta and entails a high risk for aortic dissection and rupture. These events are associated with an ultimate mortality rate of 80%. TAA is a characteristic hallmark of Loeys-Dietz syndrome (LDS), which is an autosomal dominant connective tissue disorder also presenting with multiple other skeletal, cutaneous and cardiovascular abnormalities.

A major unresolved aspect of LDS concerns the significant variability in aneurysm severity that has been observed between LDS patients. Even within the same family, the phenotype varies from completely asymptomatic to sudden death at a young age due to dissection, hinting towards the existence of genetic modifiers.

The general aim of my PhD project is to further dissect the genetic landscape of LDS by identifying genetic modifiers for SMAD3-related aortopathy. The anticipated results will immediately prove valuable for improved genetic counselling, facilitate the discovery of future personalized treatment and pave the way for preventive or curing therapies for LDS and possibly other aneurysmal diseases.

To identify potential genetic modifiers and to conclusively prove their role in LDS disease progression, we have defined three main objectives: (1) identification of SMAD3 modifiers by performing genome-wide single nucleotide polymorphism (SNP)-based linkage analysis and whole-genome sequencing (WGS), (2) creation and characterization of a SMAD3 induced pluripotent stem cell-derived vascular smooth muscle cell (iPSC-VSMC) model, and (3) CRISPR/Cas9-based genome editing and validation of the identified modifier(s) in the created and characterized SMAD3 iPSC-VSMC model.

PhD student: Joe Davis Velchev
Promotors: Bart Loeys, Aline Verstraeten & Maaike Alaerts

Towards patient-specific aorta-on-a-chip models for thoracic aortic aneurysm and dissection.

Thoracic aortic aneurysm (TAA) denotes a progressive enlargement of the thoracic aorta, entailing a significant risk for life-threatening aortic dissection and/or rupture. At present, mouse models are often used to investigate and therapeutically target the molecular defects underlying TAA, as native aortic samples of patients and, especially, control individuals are hard to collect. Yet, murine in vivo studies are often lengthy and drug testing results did previously not always recapitulate in patients. With the advent of induced pluripotent stem cells (iPSCs), the field is closing in on apt solutions to faithfully model patient and control aortas in a dish. The currently available vascular smooth muscle cell (VSMC) or endothelial cell (EC) monocultures are still overly simplified, as they fail to adequately replicate the complex multilayered and multicellular structure of the aorta. Taking advantage of available iPSCs from syndromic TAA patients (FBN1 & IPO8), my project aims to 1) develop and consolidate the validity of the first iPSC-derived TAA aorta-on-a-chip models, comprising the two VSMC subtypes populating the native ascending aorta along with a layer of arterial ECs, and 2) use the established model to further investigate the disease mechanisms underlying the relatively unexplored IPO8 syndrome. The anticipated outcomes will contribute to the replacement of mouse models (3R principle) and expedite pathophysiological TAA research and drug discovery.

PhD student: Ivanna Fedoryshchenko
Promotors: Aline Verstraeten, Bart Loeys, Ilse Luyck

Using human iPSC-derived models to investigate the divergent pathomechanisms underlying biglycan-related Meester-Loeys syndrome and X-linked spondyloepimetaphyseal dysplasia.

Pathogenic variants in biglycan cause two divergent phenotypes: Meester-Loeys syndrome (MRLS) and X-linked spondyloepimetaphyseal dysplasia (SEMDX). The latter is characterized by a disproportionate short stature and caused by missense variants. MRLS, on the other hand, is a syndromic form of thoracic aortic aneurysm that is caused by loss-of-function variants. Intriguingly, MRLS patients with partial biglycan deletions present with a more severe skeletal phenotype. To date, discriminative pathomechanisms explaining why certain biglycan mutations cause MRLS and others SEMDX remain elusive. This PhD project aims to answer this research question using induced pluripotent stem cells (iPSCs) of both patient groups and their respective (isogenic) controls. IPSC-based disease modeling provides a unique opportunity for pathomechanistic investigation in a patient-, variant- and cell type-specific manner. After the creation of disease-relevant patient-derived iPSC-vascular smooth muscle cells and -chondrocytes, I will identify cell type-specific differences between MRLS and SEMDX using (1) functional assays tailored to existing pathomechanistic insights, and (2) hypothesis-free transcriptomic and proteomic approaches. Finally, I will investigate the mutational effect of partial biglycan deletions to establish a specific MRLS genotype-phenotype association.

PhD student: Anne Hebert
Promotors: Bart Loeys, Aline Verstraeten & Josephina Meester