Direct Stimulatory Effect of PCSK9 on ApoB Expression and Secretion in Human Enterocytes. (A) The human recombinant human PCSK9 used in all experiments was characterized via immunoblotting. A 2 μg quantity of recombinant human PCSK9 was immunoblotted using 10% SDS gels and a polyclonal antibody against human PCSK9. The results confirmed the expected 62 kDa size of recombinant human PCSK9. (B) PCSK9 treatment (10 μg/mL, 24 hours) significantly stimulated the cellular expression and secretion of apoB48 protein (by 40–50%) in optimally polarized CaCo-2 cells, compared with control untreated cells, maintained in 1% FBS. (C) PCSK9 (10 μg/mL, 24 hours) markedly stimulated the cellular expression and secretion of apoB100 protein (by 40–50%) in CaCo-2 cells, compared with control untreated cells, maintained in 1% FBS. (D) The stimulatory effect of PCSK9 (24 hours) on apoB48 and apoB100 secretion in CaCo-2 cells (compared with control untreated cells maintained in 1% FBS) occurred in a dose-responsive manner in the 0 to 12.5 μg/mL concentration range of PCSK9. The maximum stimulatory effect was reached at 10 μg/mL PCSK9. (E) Time course effects of PCSK9 treated CaCo-2 cells (10 μg/mL) showed an enhancement in both apoB48 and apoB100 secretion as early as 4 hours post-treatment (versus control untreated cells) and continued on to the 24-hour time-frame of the study.