Single cell-guided prenatal derivation of primary epithelial organoids from the human amniotic fluid to model fetal tissue development and congenital disease.

Despite the advances in prenatal diagnostics, it is still difficult to predict severity and outcomes of many congenital conditions.

New patient-specific prenatal disease models are therefore needed to refine diagnosis and conduct personalised drug testing.

This AMS project aims at building a comprehensive atlas of the cellular content of the human amniotic fluid (AF) and its changes along human development.

Recent work from my team conducted using single cell RNA sequencing, showed presence in the fetal fluids of previously unobserved epithelial stem cells of different fetal tissues.

Furthermore, when put in permissive conditions, these cells formed clonal epithelial organoids manifesting fetal intestine, kidney and lung identity.

Upon specific differentiation protocols, these Amniotic Fluid-derived Organoids (AFO) showed acquisition of tissue-specific maturation features such as, in the case of lung AFOs, developing functional cilia.

With relevance for prenatal modelling, when derived from the AF of diseased pregnancies (e.g, Congenital diaphragmatic hernia, Spina bifida), AFOs manifest detectable phenotypic and functional alterations that recapitulate some pathological features of the diseases.

Building on our preliminarily dataset, this AMS proposal aims to investigate the possibility of: 1) isolating AF epithelial stem cells and AFOs from a broader gestational age range (12-40weeks) and investigate the possibility of deriving AFOs of other tissues 2) isolate and study AFOs from multiple congenital conditions to conduct phenotypic, transcriptomic and functional comparison with healthy controls.

Ultimately, this project will pave the way for the development of a patient-specific in vitro platform, to model congenital conditions and investigate personalised therapeutic interventions.