PhD defence A. (Almira) Zada

Below is a brief summary of the dissertation: 

This thesis focuses on the basic and translational research of primary pediatric intestinal pseudo-obstruction (PIPO), which results from defects in enteric nervous system (ENS) development (neuropathic PIPO) and smooth muscle function (myopathic PIPO). In Chapter 2, we identified TFAP2B as a novel candidate gene for neuropathic PIPO through whole exome sequencing (WES). A heterozygous 10-base pair deletion in TFAP2B led to exon skipping and a premature stop codon, confirming its pathogenicity. Functional assays in zebrafish demonstrated reduced enteric neurons and delayed intestinal transit, supporting TFAP2B’s role in gastrointestinal (GI) development. Chapter 3 explored Filamin A (FLNA), implicated in myopathic PIPO. We found that pathogenic FLNA variants disrupt the expression of its long isoform, essential for intestinal smooth muscle function. Zebrafish models lacking this isoform exhibited shortened intestines, hypertrophic smooth muscle, and impaired motility, linking FLNA mutations to congenital short bowel syndrome and myopathic PIPO. In Chapter 4, we investigated DNA methylation in Hirschsprung disease (HSCR), the most common neuropathic PIPO. Genome-wide methylation analysis revealed hypermethylation in ENS-related genes, particularly MAB21L2. Zebrafish models suggested an epistatic interaction between RET and MAB21L2, further implicating methylation in HSCR pathogenesis. Chapter 5 employed patient-derived induced pluripotent stem cells (iPSCs) to model HSCR. Variants in RET, GFRA1, ZEB2, and EDNRB/EDN3 affected differentiation, migration, and proliferation of enteric neuronal progenitors. Our findings highlight the potential of iPSCs in disease modeling and cell therapy strategies. Overall, this thesis provides new insights into PIPO etiology by identifying causative genes and elucidating underlying cellular and molecular mechanisms.