Soutenance de thèse Méryl ROUDAUT

Human induced pluripotent stem cell (hiPSCs)  offer an attractive alternative to study novel functions linked to diseases  and to setup innovative models. In this context, we studied a key player of the hepatic cholesterol metabolism regulation, PCSK9. Upon hiPSCs differentiation into hepatocyte-like cells, we found an unexpected role of PCSK9 in undifferentiated cells. Using tools such as overexpression, CRISPR/Cas9-mediated gene invalidation, and hiPSC derived from a patient carrying PCSK9 loss of functions mutations, we found a regulatory effect of PCSK9 on the TGFß pathway. This effect is mediated by DACT2, a negative regulator of the TGFßR subunit R1. Through DACT2 modulation by PCSK9, the SMAD2 phosphorylation is impacted, thus hiPSCs proliferation.
In parallel, to enhance hepatic cells functions generated from hiPSCs, we setup a new simplified procedure to obtain liver organoïdes. hiPSCs are cultured in a modified hydroscaffold,  BIOMIMESYS®, produced by HCS Pharma in a 96-well format suitable for molecular screening. Our procedure, recapitulating key steps of liver development, allowed us to generate liver organoids including not only hepatocytes but also, biliary-, stellate- and endothelial-cells. Functional characterizations showed enhanced cytochrome activities compared to HLCs and excellent pharmacological responses with lipid accumulation upon amiodarone or ethanol treatments and LDL-bodiopy uptake upon statin treatments. As our model can be personalized and automatized, if offers a new perspective for high content molecular screening.