Effect Of Quercetin On Regulation Of Homocysteine Metabolism And Its Mechanism

ObjectiveThe animal model of hyperhomocysteinemia was established and the effects of quercetin on homocysteine (Hey) metabolism and its molecular mechanism were explored to provide a theoretical basis for application in prevention and treatment of cardiovascular disease.Methods1. The rats were fed AIN-93diet supplemented with10g/kg,20g/kg and30g/kg methionine (Met) for21days, and serum Hey and its related metabolites and amino were determined by high performance liquid chromatography(HPLC), and amino acid analyzer repectively. The optimal dose of Met to induce hyperhomocysteinemia was investigated.2. The rats were fed AIN-93diet or supplemented with1%Met in combination with restraint stress by pair feding for6weeks. HPLC determination of serum Hey, cysteine (Cys), glutathione (GSII), and other amino acids levels were dctcrmined by HPLC and amino acid analyzer repectively.3. The rats were fed AIN-93diet supplemented with10g/kg Met and1g/kg,5g/kg and25g/kg quercetin respectively for6weeks. Hey, Cys, GSH, quercetin and its metabolites in serum were determined by HPLC method. Serine, taurine cystine and glycine were determined by amino acid analyzer. Simultaneously the liver tissue antioxidant capacity was determined. The effective dose of quercetin on regulation of Hey metabolism was established.4. The rats were fed AIN-93diet supplemented with10g/kg Met and5g/kg quercetin for6weeks. The levels of Hey, Cys, GSH in serum, and S-adenosylmethionine (SAH), S-adenosylhomocysteine (SAM) contents and S-adenosylhomocysteine hydrolase (SAHH) activities in liver were determined by HPLC. Serine, taurine, cystine and glycine were determined by amino acid analyzer., SAHH cystathionine-beta-synthase (CBS), betaine homocysteine methytransferase (BHMT), methionine synthase (MS), cystathionine-γ-lyase (CSE) activities and their mRNA were measured biochemically.Results1. The rats of hyperhomocysteinemia moled was establised2. Supplementation of1%Met to diet could increase the Hey levels of rats significantly (P<0.05) and no growth inhibition was observed. The levels of Hey in serum were significantly increased in2%and3%Met goups (P<0.05), but reduced food intake and growth inhibition were observed. No significant difference in serum Cys and GSH content in Met diet groups as compared to the control group.3. The serum Hey level in1%Met group increased according to restraint time and was significantly higher than other goups at6th week. At the end of6weeks, Heylevel in the restraint stress group (RS) was significantly lower than the control and1%Met groups(P<0.05). Serum Cys levels in RS and1%Met+RS groups were also significantly lower than the control and1%Met groups(P<0.05). The GSH level of RS rats in serum was significantly lower than other groups(P<0.05).4. The contents of Thr, Ser, Gly, Glu, Cys, Tyr, Phe, Lys and Arg in serum were signifficantly decreased (P<0.05) in RS goups, and the contents of Met and Tau were signiffiantly increased as compared with the control (P<0.05)。5. The serum contents of quercetin and isorhamnetin in rats were correlated positively to the dose of quercetin supplemented in the feed. The content of serum Hey was increased significantly after1.0%Met supplementation and reduced significantly in response to1%Met/0.5%Q group (P<0.05). The content of serum Tau was also increased upon1.0%Met supplementation. The serum contents of Ser and GSH were decreased in the quercetin supplemented groups. However, the reduction was significant only in the2.5%quercetin supplemented group as compared to the1.0%Met group (P<0.05). No significant change was found for serum content of Cys after Met or quercetin treatments. The content of serum Met was decreased significantly only in1.0%Met/2.5%Q group as compared to1%Met group without Q supplementation.6. No significant change was observed for hepatic ferric reducing ability of plasma (FRAP) value after Met and quercetin supplementations. The superoxide dismutase (SOD) activity was significantly lower in the1.0%Met/2.5%Q supplemented group than in other groups (P<0.05). The serum GSH content in three quercetin supplemented groups was significantly decreased compared to the control and1.0%Met supplemented groups (P<0.05). The content of liver malondialdehyde (MDA) in1.0%Met/0.5%quercetin supplemented group was significantly decreased compared to the control,1.0%Met,1.0%Met/2.5%quercetin supplemented groups (P<0.05). The content of carbonyls was decreased in the1.0%Met/0.5%quercetin group and increased in the1.0%Met/2.5%quercetin group, but without statistical significance.7. Serum alanine aminotransferase (ALT) and aspartate transaminase (AST) activities were significantly higher in the1.0%Met/2.5%quercetin supplemented group than in other groups (P<0.05). No significant difference was found among the control,1.0%Met,1.0%Met/0.1%quercetin,1.0%Me/0.5%quercetin groups in serum ALT and AST activities.8. Liver SAM level was significantly higher in1%Met/0.5%quercetin supplemented group than in control and1%Met groups (P<0.05). Liver SAH level was also significantly higher in1%Met/0.5%quercetin supplemented group than in control and0.5%quercetin groups (P<0.05). SAM/SAH was also significantly lower in1%Met/0.5%quercetin supplemented group than in Control,0.5quercetin and1%Met groups (P<0.05). SAHH activity was significantly increased in Met and quercetin supplemented groups (P<0.05). Compared with the Control and0.5%quercetin groups, mRNA expression of SAHH in the liver was up-regulated in1%Met and1%Met/0.5%quercetin groups (P<0.05).9. Compared with the control group, BHMT activity was significantly increased in1%Met/0.5%Q group. MS activity was significantly higher in1%Met/0.5%Q group than in other three groups. Compared with the control and0.5%quercetin groups, mRNA expression of MS in the liver was up-regulated in1%Met and1%Met/0.5%Q groups (P<0.05). Liver CBS activity was significantly increased in control,1%Met and0.5%quercetin groups, mRNA expression of CBS in the liver was up-regulated in1%Met+0.5%Q group (P<0.05). Compared with control group, mRNA expressions and activity of CSE in the0.5%Q,1%Met and Met+0.5%Q groups were significantly increased (P<0.05).Conclusion1.1%Met diet feeding could successfully induce Hey model.2. Serum Hey level was reduced and some amino acids levels also changed after restraint stress. 3. Quercetin supplementation to1%Met diet was effective in attenuating the increase of serum Hcy level in the rats. The optimal dose of quercetin acted as an antioxidant, but excess intake of quercetin was not beneficial in terms of antioxidant status in1.0%methionine induced Hcy model of the rats.4. Quercetin decreased Hcy level in serum by increasing CBS and CSE activities and mRNA expressions involved in the transsulfuration pathway.