# Effects of Bilberry Extract on Hepatic Cholesterol Metabolism in HepGG2 Cells
## 4 Discussion
Hepatic LDLR regulates the uptake of apoB- and apoE-containing lipoproteins [55]. There was a marked induction of LDLR protein levels without changes in mRNA abundance in BE-treated HepG2 cells. Furthermore, LDLR induction by BE was not dependent on SREBP2 transcriptional regulation. These results indicate that BE increased LDLR by post-transcriptional regulation mechanisms. PCSK9, the negative post-transcriptional regulator of LDLR, binds and regulates the lysosomal degradation of LDLR [56; 57]. Therefore, we examined the mRNA and protein expressions of PCSK9 to address underlying mechanisms of noticeable upregulation of LDLR protein by BE. Both PCSK9 mRNA and protein levels were significantly abolished in BE-treated HepG2 cells. A black raspberry extract containing flavonoids and phenolic acids exerted upregulation of LDLR and downregulation of PCSK9 in HepG2 cells [58]. LRP1 and SR-B1, the central regulators of plasma lipid concentration, play critical roles in the uptake of chylomicron remnant and HDL, respectively. BE treatment upregulated the expression of LRPI, whereas the expression of SR-B1 was downregulated. Polyphenol-rich compounds such as red wine concentrate, propolis and quercetin altered the hepatic SR-B1 expression in HepG2 cells and mice [59; 60; 61]. MTTP and ACAT2 play an essential role in the formation and secretion of the hepatic apoB-containing VLDL [62]. Hepatic MTTP, a lipid transfer protein, transfers TG and cholesterol esters for VLDL assembly and secretion [63]. ACAT2 catalyzes the esterification reaction that converts intracellular free cholesterol to cholesterol ester assembled into VLDL particles for secretion in the liver [64]. Dietary factors that suppress the secretion of VLDLs can be candidates for the prevention of dyslipidemia. The expression of both MTTP and ACAT2 was significantly decreased by BE. Consistent with our results, polyphenol-rich foods decreased apoB secretion attributed to the reduction in MTTP in HepG2 cells. Ellagic acid, a natural polyphenic compound, suppressed apoB secretion in HepG2 cells [65]. White tea extract rich in EGCG downregulated apoB and MTTP expression, followed by a reduction in VLDL production [66]. Hepatic biliary cholesterol efflux is governed by canalicular cholesterol transporters, such as NPC1L1 and the heterodimer ABCG5/G8 [67]. Highly expressed in the intestine and liver, NPC1L1 functions as a cholesterol absorption transporter in humans [68]. One of the commonly prescribed medicine to lower plasma cholesterol is ezetimibe, the NPC1L1 inhibitor that blocks intestinal cholesterol absorption [69; 70]. Furthermore, NPC1L1 in hepatocyte counterbalances biliary cholesterol secretion mediated by ABCG5/G8 [71]. The hepatic ABCG5/G8 facilitates the direct export of free cholesterol from the liver to bile. The mRNA and protein levels of NPC1L1 were reduced, whereas ABCG8 was markedly elevated in BE-treated HepG2 cells. The NPC1L1 expression was directly and indirectly inhibited by luteolin and phenolic-compound-rich _Opuntia focus indica_[72; 73]. Similar to this study, persimmon tannin and coffee arabica pulp increased the expression of ABCG5/G8 in HepG2 cells [74; 75]. Bile acid conversion from cholesterol is an essential pathway for cholesterol elimination. CYP7A1 and ABCB11 are required for biliary cholesterol excretion. In the liver, CYP7A1 initiates cholesterol degradation by converting cholesterol to bile acids [76]. The bile salt export pump (BSEP), ABCB11, facilitates canalicular bile acid secretion [77]. There was a marked upregulation of genes for rate-limiting control of bile acid biosynthesis and secretion in BE-treated HepG2 cells. Polyphenols, i.e., resveratrol and quercetin, induced CYP7A1 expression in HepG2 cells [78]. All of the genes for cholesterol transport were changed in the direction of canalicular biliary cholesterol secretion in BE-treated HepG2 cells.