Fig. 6. PAI-1 deficiency leads to lower concentrations of plasma apoB and apoB-cholesterol in mice and humans.

(A) Serpine1^fl/fl mice were treated with AAV8-TBG-Cre (Cre) or control AAV8-TBG-LacZ (Ctrl) and then fed a high-fat diet for 8 weeks. Plasma total cholesterol concentration was measured (n = 6 per group). (B) Serpine1^fl/fl mice were treated with AAV8-TBG-Cre (Cre) or control AAV8-TBG-LacZ (Ctrl) and then fed a high-fat diet for 8 weeks. Plasma apoB was measured by immunoblot (n = 6 per group). (C) Serpine1^fl/fl mice were treated with AAV8-TBG-Cre (Cre) or control AAV8-TBG-LacZ (Ctrl) and then fed a high-fat diet for 8 weeks. Plasma samples were subjected to FPLC fractionation and assayed for cholesterol concentration. The cholesterol in VLDL fractions is shown in the smaller graph in the inset (n = 6 per group). (D) Serpine1^fl/fl mice were treated with AAV8-TBG-Cre (Cre) or control AAV8-TBG-LacZ (Ctrl) and then fed a high-fat diet for 8 weeks. Liver lysates were assayed for PAI-1–free tPA concentration by ELISA (n = 6 per group). (E) Serpine1^fl/fl mice were treated with AAV8-TBG-Cre (Cre) or control AAV8-TBG-GFP (Ctrl) and then fed a normal chow diet for 4 weeks. Mice were injected with P407 i.p. to assess VLDL secretion (n = 10 per group). (F) Serpine1^fl/fl mice were treated with AAV8-TBG-Cre (Cre) or control AAV8-TBG-GFP (Ctrl) and then fed a normal chow diet for 6 weeks. The mice had their food withdrawn for 5 hours and then were euthanized at 0, 2, and 4 hours after oral gavage with olive oil. Plasma apoB-100 concentration was measured by ELISA. (G) Plasma samples from SERPINE1-deficient humans and unaffected age-, gender-, and BMI-matched individuals from the same community were assayed for VLDL cholesterol, LDL cholesterol, and apoB-100 concentrations (n = 10 per group). (H) tPA concentration was measured in VLDL isolated by ultracentrifugation from the plasma of the subjects in (G) (n = 10 per group). (I) VLDL from the plasma of the subjects in (G) was analyzed by dynamic light scattering (n = 10 per group). The correlation between VLDL-associated tPA and VLDL diameter was calculated (n = 10 per group). (J) A schematic diagram depicting how tPA–PAI-1 interaction in hepatocytes determines VLDL assembly. Without lipid stimulation, tPA interacts with apoB and inhibits MTP-apoB interaction in the ER, thereby limiting MTP-mediated apoB lipidation and VLDL assembly. When hepatocytes are loaded with lipid, PAI-1 sequesters free tPA away from apoB and increases VLDL assembly. The diagram was generated using biorender.com. Data are shown as means ± SEMs; P values were calculated by two-tailed Student’s t test [(A), (D), (E), and (F)], paired Student’s t test [(G) and (H)], or Pearson’s correlation analysis (I). *P < 0.05.