Regulation Effects Of Quercetin On Cholesterol Metabolism

Objective Based on previous works, several receptor proteins, rate-limiting enzymes and transcriptional regulation factors involved in cholesterol metabolism were analyzed at m RNA or protein expression level in rats after quercetin supplementation so as to explore the mechanisms whereby quercetin regulates cholesterol metabolism and to provide an experimental basis for understanding the roles played by quercetin in vivo.Materials and Methods 1. Influence of quercetin on blood lipids and cholesterol content in arcus aortae. Twenty four healthy male Wistar rats were divided randomly into control group and quercetin supplemented group based on body weight. Control group was fed AIN-93 G diet. Quercetin supplemented group were fed the AIN-93 G diet supplemented with 0.4% quercetin. The rats were housed and fed individually for 5 weeks. Dietary intake was recorded daily and body weight was monitored weekly. Blood samples were taken from posterior orbital venous plexus at weeks 2, 4 and 5 and the levels of serum TC(total cholesterol), TG(triacylglycerol), LDL-C(low-density lipoprotein cholesterol) and HDL-C(high-density lipoprotein cholesterol) were determined. At the end of the experiment, the rats were sacrificed under ether anesthesia and arcus aortae were separated to measure the contents of TC, FCH, and CE. 2. Influence of quercetin on LDLR expression The rats were sacrificed after 5 weeks feeding and the liver and arcus aortaes were taken for examining the expression of LDLR(low-density lipoprotein receptor), which is involved in the process of LDL-C uptake. Real-time PCR method was applied to determine LDLR gene expression and Western blotting method to determine LDLR protein expression in the liver and arcus aortae. 3. Influence of quercetin on the expression of cholesterol transporters Real-time PCR method was used to measure ABCA1(ATP-binding cassette transporter A1), ABCG1(ATP-binding cassette transporter G1), LXRα(liver X receptor α) m RNA expression and Western blotting method to measure ABCA1, ABCG1 and LXRα protein expression in the liver and arcus aortae. 4. Influence of quercetin on hepatic cholesterol synthesis The liver microsome was extracted by ultra-centrifugation and an ELISA assay was used to measure the HMG CR activity in the liver microsome. Real-time PCR method was applied to detect the mRNA expression of HMG CR and SREBP-2(sterol regulatory element-binding protein-2). Western blotting method was used to examine the protein expression of HMG CR and SREBP-2. 5. Influence of quercetin on hepatic cholesterol conversion The liver was used for measuring CYP7A1(cholesterol 7α-hydroxylase) activity, m RNA and protein expression. The liver microsome was extracted by ultracentrifugation. The endogenous cholesterol was catalyzed to generate 7α-hydroxycholesterol, which is eventually turned into HCO(7α-hydroxy-4-cholesten-3-one by HPLC). The activity of CYP7A1 was determined as the amount of HCO produced. Real-time PCR method was applied to monitor the m RNA expression of CP7A1, FXR(farnesoid X receptor) and LXRα. Western blotting method was employed to measure the protein expression of CYP7A1, FXR and LXRα.Results 1. Influence of quercetin on blood lipids and cholesterol content in arcus aortae There was no difference in dietary intake and body weight between the control group and the quercetin supplemented group(P>0.05).Serum LDL-C concentration in rats exposed to 0.4% quercetin was significantly higher than those in the control group(P<0.05) after 5 weeks feeding. However, serum TC、TG and HDL-C showed no statistical difference between the two groups(P>0.05). Compared with the control group, the contents of TC,FCH and CE in arcus aortae declined obviously in rats treated with 0.4% quercetin(P<0.05). 2. Influence of quercetin on LDLR expression After 5 weeks of 0.4% quercetin treatment, LDLR m RNA expression was significantly increased in both liver and arcus aortae(P<0.05). Meanwhile, LDLR protein expression rose significantly(P<0.05), indicating that more LDL-C can be uptaken from the circulation. 3. Influence of quercetin on the expression of cholesterol transporter proteins Treatment with 0.4% quercetin for 5 weeks obviously increased the m RNA expression of ABCA1 and ABCG1 in the arcus aortae, whereas the m RNA expression of LXRα was also increased significantly(P<0.05). In the liver, ABCA1 expression showed no statistical differences at m RNA and protein levels between the two groups(P>0.05). However, hepatic ABCG1 expression was higher in the quercetin supplemented group than in the control group(P<0.05). At the same time, LXRα expression at the m RNA and protein levels also rose significantly(P<0.05). 4. Influence of quercetin on hepatic cholesterol synthesis It was showed that HMG CR activity displayed no statistical difference between the quercetin supplemented group and the control group(P>0.05), though a declining tendency was observed. However, HMG CR m RNA expression decreased obviously compared with the control group(P<0.05). HMG CR protein expression showed no statistical difference(P>0.05) between the two groups. On the other hand, no statistical difference was noted for SREBP-2 expression at both m RNA and protein levels(P>0.05). 5. Influence of quercetin on hepatic cholesterol conversion In the quercetin supplemented group, hepatic CYP7A1 activity rose significantly compared with the control group(P<0.05). Its expression at both m RNA and protein levels also was increased significantly(P<0.05). No statistical difference was found for FXR expression at the m RNA and protein levels between the two groups(P>0.05). However, LXRα expressed significantly higher at both m RNA and protein levels after quercetin treatment(P<0.05).Conclusion 1. It is indicated that quercetin can affect the cholesterol metabolism significantly by changing the distribution of cholesterol, decreasing the content of TC, FCH and CE in the arcus aortae and other tissues, and increasing the concentration of LDL-C in serum. 2. Treatment with 0.4% quercetin can significantly increase the expression of LDLR at both m RNA and protein levels in the liver and arcus aortae, which promotes the uptake of circulating LDL-C. 3. Quercetin can promote the m RNA expression of ABCA1 and ABCG1 possibly via increasing the m RNA expression of LXRα in the arcus aortae. It also can promote the expression of ABCG1 at both m RNA and protein levels via increasing the expression of LXRα at both m RNA and protein levels, indicating that quercetin can increase the cellular efflux of cholesterol and decrease the content of cholesterol in the peripheral tissues. 4. Based on the results of the activity, gene or protein expression of HMG CR, it is concluded that quercetin has limited impact on the cholesterol synthesis. 5. Quercetin can increase the activity, gene or protein expression of CYP7A1, suggesting that quercetin can promote the efflux of cholesterol by promoting the conversion of cholesterol to bile acids. This action is mediated partially by increasing the expression of LXRα at both m RNA and protein levels..