White_2025_bioRxiv__

Current lipid-lowering drugs are relatively ineffective in reducing low-density lipoprotein (LDL) cholesterol in patients with Familial Hypercholesterolemia (FH) due to a dysfunctional LDL receptor (LDLR). However, LDL cholesterol reductions have been achieved in FH patients using angiopoietin-like 3 (ANGPTL3) inhibitors, which act through an uncharacterized, LDLR-independent pathway that requires endothelial lipase (EL). Here, we aim to investigate EL's direct role in LDLR-independent uptake of LDL in hepatocytes. Control and LDLR-KO HepG2 cells were transfected with an empty plasmid or a plasmid encoding the human LIPG gene, and the cellular uptake of fluorescent human LDL was measured by FACS. To test the contribution of heparan sulfate proteoglycans (HSPG), LDL uptake was assessed with and without the pre-incubation with heparin or a heparinase cocktail. Finally, tetrahydrolipstatin was used to inhibit EL enzymatic activity in uptake studies. Unsurprisingly, LDLR-KO HepG2 cells showed an 80% reduction in LDL uptake compared to controls (p<0.001). Remarkably, EL overexpression almost fully rescued LDL uptake in LDLR-KO cells (p<0.001), without effects in control cells. EL-mediated LDL uptake was completely blocked by heparinases and heparin in LDLR-KO cells, suggesting a crucial role of HSPG in EL-mediated LDL uptake. Notably, treatment with tetrahydrolipstatin reduced LDL uptake in EL-overexpressing LDLR-KO cells (p=0.0015). Our data reveals that EL facilitates the uptake of LDL in hepatocytes through an LDLR-independent, HSPG-dependent pathway that involves EL's enzymatic activity. This pathway provides an additional mechanism to explain the reduction of LDL cholesterol induced by ANGPTL3 inhibitors and represents a potential druggable target to treat FH.