Hepatocyte Familial Hypercholesterolemia (FH)

DefiniGEN provide disease modelled familial hypercholesterolemia hepatocytes which are highly functional CRISPR gene-edited human hepatocytes. Familial hypercholesterolemia (FH) is an autosomal disorder of lipoprotein metabolism caused mainly by mutations in the low-density lipoprotein receptor (LDLR) gene. These cells can offer disease modelling and drug discovery researchers unprecedented tool for elucidating the underlying mechanisms of this disease.

• Highly standardized cell product containing >98% human hepatocyte cells with consistent performance and biologically relevant data

• Disease circuit verified mutation in the LDLR gene E101K results in significantly impaired LDL uptake in the cells

Genetic validation and cell viability

Def-HEP FH cells have been validated and verified for the E101K genetic mutation in the LDLR gene. Typical viabilities of the thawed hepatocytes are >70% upon receipt.

Disease circuit verification

Figure 1. Sequence confirmation of LDLR E101K mutation in Def-HEP FH cells. Verifying the correct heterozygous C.301CG>A substitution in the LDLR gene.

Functional test

The in vivo functional implications of LDL receptor deficiency are conserved in our model, as shown by immunostaining and FACS analysis demonstrating that Def-HEP FH iPSC hepatocytes have an impaired ability to incorporate LDL receptor-specific binding of Dil-LDL is followed by internalization of the bound complex and lysosomal hydrolysis of the ligand. Increase in the fluorescence intensity per cell is hence used as a measure of Dil-LDL uptake and, implicitly, as an indication of LDL-R presence. These results demonstrate that CRISPR generated disease-specific human iPSC can successfully be used to model FH.

LDL receptor analysis

Def-HEP FH cells demonstrate an impaired ability to take up LDL cholesterol relative to the WT isogenic control.

Figure 3. Quantitative LDL receptor assay. Receptor assays based on fluorescent Dil-LDL internalization have demonstrated that Def-HEP FH iPSC-derived hepatocytes have a significantly impaired ability to incorporate LDL relative to the isogenic control Def-HEP WT over a range of LDL substrate ranges.