Mechanism Of Chlorogenic Acid On Apoptosis In HepG2 Cells

Since the 1990s, hepatoma has been ranked the second mortality in china. Chlorogenic acid is one of the most abundant polyphenolic compounds in human diet, and it is also well known for its beneficial biological properties, such as anti-oxidant, anti-bacterial, and anti-cancer effects. In this paper, the mechanism of apoptosis in human hepatocellular carcinoma cell line HepG2 induced by chlorogenic acid was studied by two apoptotic signaling pathways, which are instrinsic pathway and extrinsic pathway.Human hepatocellular carcinoma cell line HepG2 were cultured in vitro, and Human liver cells L02 were cultured as normal control. The influence of chlorogenic acid in different concentrations on the proliferation of HepG2 and L02 were detected by the method of CCK-8, and then analyze the IC50 value through SPSS software, obtain the optimum experiment conditions. Lactate dehydrogenase activity in cell culture supernatant was examined by lactate dehydrogenase kit, flow cytometry was used to detect the apoptosis rates, effect of chlorogenic acid on apoptosis related genes and protein expression was analysed by qRT-PCR and Western blot.CCK-8 assay results showed that chlorogenic acid inhibited HepG2 cells proliferation, and the inhibition of chlorogenic aid on L02 cells was unconspicuous. The IC50 of chlorogenic acid after 24 h to 72 h were 212.04±7.15 ?M?22.17±1.13?M?18.58±1.07 ?M, compared with IC5048h, IC5024h and IC5072h were significantly difference(P< 0.01), but there was no difference between IC5072h and IC5048h. Lactate dehydrogenase activity in the cell culture supernatant of HepG2 cells was significantly increased after treated by chlorogenic acid after 48 h compared with normal control group (P< 0.01), the difference in L02 cells was not obvious. The result of flow cytometry showed that apoptosis rates of HepG2 normal control group?HepG2 CGA group?HepG2 5-Fu group?L02 normal control group?L02 CGA group were 1.85%?35.98%?20.1%?1.87%?4.7%. Compared with HepG2 NC group, the apoptosis rates of HepG2 group and HepG2 5-Fu group were statistically significant (P< 0.01), the apoptosis rates of HepG2 drug groups were also increased significantly compared with L02 CGA group (P< 0.01). The result of qRT-PCR showed that chlorogenic acid can promote the expression of Fas?FasL?FADD?Caspase 3? Caspase 8?Caspase 9?Bim?Bak? Bid and inhibited the expression of Bcl-xL at the same time. The result of Western blot showed that chlorogenic acid promote the release of p53? Cyto c?Smac through lowering the Bcl-2/Bax ratio, and increased the expression of Bid and Apaf-1 protein, decreased the expression of cIAP-1 to induce HepG2 cells apoptosis.Chlorogenic acid induced HepG2 cells apoptosis through increasing the expression of pro-apoptotic factors and decreasing the expression of anti-apoptotic factors.