Transforming growth factor beta reduces proprotein convertase subtilisin/kexin type 9 in the supernatant of hepatic stellate cells

Figure 1 Proprotein convertase subtilisin/kexin type 9 in the supernatants of 10⁶ liver cells cultivated for 24 hours, cellular levels of HepG2, Huh7, and LX-2 cells, and of LX-2 cells during culture. A: Proprotein convertase subtilisin/kexin type 9 (PCSK9) in the supernatant of LX-2 cells (6 different cultures), primary human hepatic stellate cells from 9 donors, primary human hepatocytes from 10 donors, Huh7 cells (16 different cultures), HepG2 cells (5 different cultures), human hepatic sinusoidal endothelial cells from 3 donors and primary human Kupffer cells from 2 donors; B: Immunoblot of PCSK9 protein in Huh7 and HepG2 cells, glyceraldehyde-3-phosphate dehydrogenase was used for normalization; C: Immunoblot of PCSK9 protein in LX-2 and HepG2 cells, cyclophilin A was used for normalization; D: PCSK9 protein in the supernatant of LX-2 cells cultivated for 24 hours, 48 hours, and 72 hours. PCSK9: Proprotein convertase subtilisin/kexin type 9; HSC: Hepatic stellate cell; PHH: Primary human hepatic; HHSEC: Human hepatic sinusoidal endothelial cell; KC: Kupffer cell; GAPDH: Glyceraldehyde-3-phosphate dehydrogenase.

Figure 2 Effect of lipopolysaccharide and transforming growth factor beta on proprotein convertase subtilisin/kexin type 9 in the culture medium of LX-2 cells. A: Proprotein convertase subtilisin/kexin type 9 (PCSK9) in the LX-2 cell culture medium after 24 hours of cultivation with 0 μg/mL, 1 μg/mL, or 10 μg/mL lipopolysaccharide (4 independent experiments); B: PCSK9 in the LX-2 cell culture medium after 24 hours of cultivation with 0 ng/mL, 5 ng/mL, or 7 ng/mL transforming growth factor beta (TGF-β; 6 independent experiments for controls and 7 ng TGF-β, and 3 independent measures for 5 ng/mL); C: PCSK9 protein of LX-2 cells cultivated with 0 ng/mL, 5 ng/mL, or 7 ng/mL TGF-β for 24 hours. PCSK9: Proprotein convertase subtilisin/kexin type 9; TGF-β: Transforming growth factor beta; GAPDH: Glyceraldehyde-3-phosphate dehydrogenase.

Figure 3 Regulation of proprotein convertase subtilisin/kexin type 9 levels in LX-2 cells by cytokines, adipokines, and nuclear receptor agonists. A and B: Proprotein convertase subtilisin/kexin type 9 (PCSK9) levels in LX-2 cells cultivated with tumor necrosis factor and interleukin-6. PCSK9 levels in the LX-2 cell supernatant after 24 hours of cultivation with 0 ng/mL, 0.2 ng/mL, 2 ng/mL, or 4 ng/mL tumor necrosis factor, 4 independent experiments (A); PCSK9 levels in the LX-2 cell supernatant after 24 hours of cultivation with 0 ng/mL, 5 ng/mL, 10 ng/mL, or 20 ng/mL interleukin-6, 4 independent experiments (B); C-E: PCSK9 levels during LX-2 cell cultivation with adipokines and chemerin isoform overexpression. PCSK9 levels in the LX-2 cell culture supernatant after 24 hours of cultivation with 0 μg/mL or 10 μg/mL adiponectin, 6 independent experiments (C); PCSK9 levels in the LX-2 cell culture supernatant after 24 hours of cultivation with 0 ng/mL, 4 ng/mL, 10 ng/mL, or 20 ng/mL leptin, 4 independent experiments (D); PCSK9 in the supernatant of LX-2 cells cultivated for 24 hours with overexpressed chemerin-155, chemerin-156, or chemerin-157 and in control transfected cells, 4 independent experiments (E); F-H: PCSK9 levels during LX-2 cell cultivation with nuclear receptor agonists. PCSK9 levels in the LX-2 cell culture supernatant after 24 hours of cultivation with 0 μmol/L, 1 μmol/L, and 2 μmol/L GW4064, 5 independent experiments (F); PCSK9 levels in the LX-2 cell culture supernatant after 24 hours of cultivation with 0 μmol/L, 5 μmol/L, and 10 μmol/L T0901317, 3 independent experiments (G); PCSK9 in the supernatant of LX-2 cells cultivated for 24 hours with 0 μmol/L, 7.5 μmol/L, and 15 μmol/L rosiglitazone, 3 independent experiments (H). PCSK9: Proprotein convertase subtilisin/kexin type 9; TNF: Tumor necrosis factor; IL: Interleukin; Chem: Chemerin.