Effects Of Cyanidin On Liver Cell Lipid Metabolism In Vitro

Anthocyanin, also known as Anthocyanidin, is a flavonoid compound. It is a water-soluble natural pigment widely present in plants, it is also the main coloring material in petals. There are more than three hundred anthocyanins and cyanidin (CY) is the simplest chemical form of anthocyanin glycosides. All other anthocyanins contain the molecular structures of cyaniding and one part of anthocyanins can be formed in vivo during the metabolism of cyanidin. It has been found that cyanidin has different biological activities such as cancer prevention, cardiovascular and epithelial cell protection, fat metabolism regulation, and liver protection against mycotoxins. These biological activities of cyanidin are closely related to its absorption, transportation and metabolism. Prior experiments have shown that CY can improve the oxidative stress status of animals and regulate the biological activity of lipid metabolism. These data indicated that the application of CY on animal production may significantly improve animal performance and promote their resistance to diseases. However, reports regarding its pharmacological effects, pharmacokinetics and in vivo animal studies are relatively limited. In this study, we explored the effects of cyanidin on liver cell lipid metabolism in vitro which may provide theoretical basis to support the potential application of CY in livestock production. In vitro hepatic steatosis model was developed, and effects of CY on liver cell lipid metabolism, oxidative stress, lipo-apoptosis, intrahepatic lipid metabolism and in vitro signal pathway were studied.1. Selection of the optimal cell line:L02and HepG2were used in this study, results showed that low concentration of cyanidin (40μg/mL) could inhibit the HepG2cell growth but no effects was observed in L02cells which indicated that L02was more tolerable to CY than HepG2. Further detections using RT-PCR showed that all target pathway signaling designed in the present study including PPARa, PPARy, ABCA1, CYP7A1and RXRa could be detected in L02cells, but only PPARa, PPARy, ABCA1except CYP7A1and RXRa could be detected in HepG2cells. All these data indicated that L02cell is more optimal than HepG2used to develop a hepatic steatosis model.2. Establishment of the hepatic steatosis model:Palmitic acid (PA) was used to establish the hepatic steatosis model. L02cells were treated using different doses of PA,50mg/mL PA could inhibit the proliferation of the L02. Thirty mg/mL PA could induce the accumulation of triglycerides (TG) and cholesterol (TC) with dose-dependent pattern in cells. mRNA and preotein Levels of RXRa and ABCA1showed a significant increase in L02hepatic steatosis cell model, but PPARa, PPARy and CYP7A1presented a decrease. Furthermore, PA caused oxidative damage with accumulation of malondialdehyde (MDA) and reduction of hydrogen peroxide enzyme (CAT), superoxide dismutase (SOD) and glutathione (GSH) in L02cells. The intensity of the damage was also dependent on the PA dose. Moreover. Caspase-3and Caspase-9were activated and increased in L02cells after oxidative damage which resulted in cell apoptosis. Most of the apoptotic cells were limited to the early stage and the rate of cell apoptosis was positively correlated with PA dose. All above data indicated that using PA as an inducing agent (the maximum non-toxic concentration is30mg/mL) could cause abnormal liver metabolism and oxidative demage in L02cells with no effect on its vitality which means these conditions are suitable to establish the hepatic steatosis cell model..3. Effects of cyaniding on the L02hepatic steatosis model:L02hepatic steatosis model was treated with different dose of CY, data showed that CY could significantly decreased the intercellular TG and TC to improve the status of lipid accumulation in model cell. When the CY dose was≥5μg/mL, the effect was obvious. CY could down-regulated the intercellular LXRa and inhibited the expression of ABCAl in both mRNA and protein levels; CY also increased the levels of PPARa, PPARy and RXRa as well the down-stream signaling CYP7A1which promoted the metabolism of TG and TC with dose-dependent manner. Moreover, CY could increase the level of MDA and increase CAT, SOD and GSH which helped to relief the oxidative damage and inhibit the cell apoptosis through inhibition the activation of caspase-3and caspase-9in this cell model.In conclusion, the presnet study confirmed that the CY, through a variety of ways, regulated lipid metabolism, alleviated cellular oxidative damage caused by lipid accumulation and reduced fat apoptosis rate through inhibiting caspase-3and caspase-9in the L02hepatic steatosis cell model induced by PA.