Experimental Study Of Effects Of Dicaffeoylguinic Acid Against Liver Injury And Hepatofibrosis In Rats

I ,5-di-O-caffeoylguinic acid (IBE5) is an effective component extracted from Chinese herb Inula Flower, which belongs to caffeoylguinic acid compounds. It was Previously reported that cyrarine, with the similar chemical structure has beneficial effects for liver and gallbladder and can protect liver from damages induced by macrophages or CC!4. Whether IBE5 has therapeutical effect on liver disorders are interested. This paper studied the effects of IBE5 on liver both in the cellular and system levels in order to provide experiment evidence for clinical applications. Its possible pharmacological mechanism was also discussed. 1. Extraction and purification The processing of distillation and separation of Inu!a Flower includes the following steps: Inula Flower-alcohol extraction, acetic ester extraction and purification with silica gel Sephadex LH2O column. Purified IBE5 was obtained. 2 Study in vitro 2.1 Protective effects of IBE5 on cultured primary hepatocyte To study the protective effects on liver damaged by DM14 and CC!4, cultured primary rat hepatocytes were used. Hepatocytes were isolated and cultured by adding DMN and CC!4. The proliferation and the viability of the hepatocytes were evaluated by measuring the incorporation of 3H-TdR and 3H-Pro, respectively. The apoptosis and the cell cycle of the hepatocytes were examined by flow cytometer (FCM). The activity of AST was detected by AST Kit. Results showed that IBE5(7.8?50 mg.U? had no toxicity on the hepatocytes. It significantly increased the proliferation and the viability of the normal and injured hepatocytes, decreased the activity of AST in the cultural medium and restrained the hepatocytic apoptosis induced by DM14 and CCI4. In conclusion, IBE5 has notable anti-hepatocytic damage effects in vitro. 2.2 Effect of IBE5 on Proliferation and Function of Fibroblasts (NIH/3T3) To study the effects of Dicaffeoylquinic Acid (IBE5) on proliferation, viability, function of Fibroblasts, and the possible mechanism for protecting against fibrosis on liver. As the substitutive model of liver fibroblasts, Nll{13T3 fibroblasts were cultured with regular method jy~ yjfro, and incubated with 250,125,62.5,31.25,15.6,7.8 mg.L~?IBES for 24 hours. The cell viability and hylturonic acid level was assayed by [3H]-Pro incorporation, and cell proliferation by [3H]-TdR incorporation respectively. Cell collagen synthesis rate was measured with [3H]-Pro incorporation collagenase digestion. The results that the addition of IBE5 (7.8 mg.U?-- 250 mg.U? significantly reduced proliferation~ inhibited intracellular cell viability and decreased the hyaluronic acid level in a dose-dependent manner. In conclusion, IBE5 can significantly reduced proliferation,inhibited intracellular cell viability, proliferation of 313 cells, syntheses of collagen and hyaluronic acid level. 3 Study in vivo 3.1 Effects of IBE5 on liver damage induced by combined factors To study effects of IBE5 on liver fibrosis and lipid peroxidation, the rat model of hepatic fibrosis was produced by combined factors and treated with IBE5 (50 mg.kg挆200 mg.kg?. Cytolysis-marker activity in serum (ALT and AST) , serum levels of laminin (LN) hyaluronic acid (HA), lipid peroxide malondialdehyde (MDA) and nitrogen oxide (NO) were measured. The liver tissues were examined by lig...