Gene Silencing of ANGPTL3 Induces PCSK9: Exploring the Biological Significance in the Hepatoma Huh7 Cell Line

Angiopoietin-like 3 (ANGPTL3) and proprotein convertase subtilisin/kexin type 9 (PCSK9) are key regulators of lipid homeostasis. We have previously shown that gene silencing of ANGPTL3 significantly induces PCSK9 expression in the human hepatoma cell line Huh7. Here, we investigated the biological significance of this regulation in the cultured human hepatoma cell line Huh7.

We performed an RNA-seq analysis in Huh7 cells transfected with siRNA-ANGPTL3, siRNA-PCSK9, and double siRNA-ANGPTL3/PCSK9. Selected findings were assessed by RT-qPCR, Western blotting, and ELISA.

Among 13,945 detected transcripts, 192 genes were differentially expressed after ANGPTL3 silencing, 88 after PCSK9 silencing, and 219 after combined ANGPTL3/PCSK9 silencing, compared with scramble-siRNA controls. When ANGPTL3 gene expression was silenced, we observed a compensatory induction in PCSK9 mRNA and protein expression. Bioinformatic analysis revealed that gene silencing of ANGPTL3 or both ANGPTL3/PCSK9 suppresses serpin family A member 1 (SERPINA1), which encodes α1-antitrypsin, and lectin mannose-binding 1 (LMAN1). These data were confirmed by Western blot and RT-PCR analysis. In addition, ANGPTL3-siRNA, alone or combined with PCSK9-siRNA, significantly increased FV and FVIII mRNA expression and secretion in conditioned medium.

Our data identified SERPINA1 and LMAN1 as genes downregulated in response to ANGPTL3 silencing in Huh7 hepatoma cells, which was also associated with increased expression of FV and FVIII. Our study suggests a potential link between ANGPTL3 silencing and coagulation-related processes, extending the biological relevance of ANGPTL3 beyond lipid metabolism.