Abstract
Birth defects affect 3% of children in the United States. Among the birth defects, congenital heart disease and craniofacial malformations are major causes of mortality and morbidity. Unfortunately, the genetic mechanisms underlying craniocardiac malformations remain largely uncharacterized. To address this, human genomic studies are identifying sequence variations in patients, resulting in numerous candidate genes. However, the molecular mechanisms of pathogenesis for most candidate genes are unknown. Therefore, there is a need for functional analyses in rapid and efficient animal models of human disease. Here, we coupled the frog Xenopus tropicalis with Optical Coherence Tomography (OCT) to create a fast and efficient system for testing craniocardiac candidate genes. OCT can image cross-sections of microscopic structures in vivo at resolutions approaching histology. Here, we identify optimal OCT imaging planes to visualize and quantitate Xenopus heart and facial structures establishing normative data. Next we evaluate known human congenital heart diseases: cardiomyopathy and heterotaxy. Finally, we examine craniofacial defects by a known human teratogen, cyclopamine. We recapitulate human phenotypes readily and quantify the functional and structural defects. Using this approach, we can quickly test human craniocardiac candidate genes for phenocopy as a critical first step towards understanding disease mechanisms of the candidate genes.
Highlights
In order to define the genetic causes of CHDs, including craniocardiac malformations, the NIH/NHLBI established the Pediatric Cardiac Genomics Consortium (PCGC) under the Bench to Bassinet program[9]
For functional screening of craniocardiac candidate genes, we propose to model these diseases in Xenopus tropicalis, an inexpensive and rapid amphibian model, where cardiac and facial structures can be simultaneously assessed
We embedded stage 44–46 tadpoles in an agarose gel to position the ventral cardiac sac towards the optical coherence tomography (OCT) beam for imaging (Fig. 1a)
Summary
In order to define the genetic causes of CHDs, including craniocardiac malformations, the NIH/NHLBI established the Pediatric Cardiac Genomics Consortium (PCGC) under the Bench to Bassinet program[9]. The efficacy of OCT in Xenopus as well as other models is established, we currently lack a rapid, efficient and reproducible approach to imaging cardiac and facial structures in high-throughput model systems that would be amenable to candidate gene screening for craniocardiac malformations. We have not yet defined normative values for these structures nor have we demonstrated that these parameters are altered in modeled diseases To address this problem and capitalize on the benefits of OCT applied to Xenopus tadpole morphometry, we first need to establish imaging guidelines and reference data similar to clinical ultrasonography. This will provide a foundation to interrogate and quantify craniocardiac structures during normal development and when modeling disease states. Our fast and efficient approach aims to improve candidate “genes-to-functions screens” that can act as a springboard for mechanistic studies of craniocardiac malformation genes
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