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We previously showed that human pluripotent stem cells (hiPSCs) provide a suitable model to study metabolic diseases upon hepatocyte-like cell (HLC) differentiation. In particular, HLCs have been used to model cholesterol metabolism regulation, by mimicking the main disease features in vitro. Human iPSCs can be generated from urine samples of patients with a well-described phenotype and carrying specific genotypes. This non-invasive approach allowed the study of LDLR- and PCSK9-mediated autosomal dominant hypercholesterolemia (ADH) as well as PCSK9-mediated familial hypobetalipoproteinemia (FHBL). While the direct link between hiPSCs and patients, as well as the abundance of HLCs provide promising advantages of such strategy, it is impaired mainly by the neonatal characteristic of HLCs as well as the difficulty to perform high throughput studies for pharmacological investigations.
Therefore, to overcome these burdens, we choose to 1. Differentiate hiPSCs into HLCs in a 3D environment instead of the classical 2D culture systems to enhance their maturation; 2. Adapt our 3D differentiation process to a 96 wells format to make it compatible for drug screening.
To reach our goals, we established a partnership with HCS Pharma, which has an expertise in high content phenotypic screening and produces an innovative 3D scaffold, BiomimesysTM. This scaffold is composed of hyaluronic acid that can be functionalized with extra cellular matric derivatives, with adjustable stiffness and porosity. We setup conditions for hiPSCs seeding and differentiation to reach a new protocol adapted to a 3D environment. Our preliminary data indicate that our procedure enhanced expression of hepatic markers such as transcription factors (FOXA2, FOXA3, HNF1a, HNF1b, HNF4a), cytochrome P450 (CYP450) family members (CYP3A4, CYP2A6, CYP7A1) or cholesterol metabolism regulators (PCSK9, Lipoprotein(a)). During our presentation, we will discuss our data hiPSCs differentiation in 3D, CYP450 activities and induction, as well as their application for the study of metabolic diseases.