Methyl Helicterate Inhibits Hepatic Stellate Cell Activation Through Modulation of Apoptosis and Autophagy.

BACKGROUND/AIMS: Activated hepatic stellate cells (HSCs) are the major source of extracellular matrix (ECM). Therefore inhibiting HSC activation is considered as an effective strategy to inhibit the process of liver fibrosis. This study aimed to investigate the underlying mechanism of methyl helicterate (MH) isolated from Helicteres angustifolia on the activation of HSCs. METHODS: HSC-T6 cells were treated with various concentration of MH and autophagy was inhibited by 3-Methyl adenine (3-MA) or RNA interference. Cell viability was observed by MTT assay and cell colony assay. Cell cycle and apoptosis were analyzed using flow cytometry. Autophagic vacuoles were observed by transmission electron microscopy and monodansyl cadaverine (MDC) staining. Moreover, autophagy-related genes and proteins were detected using real-time PCR and Western blot assays, respectively. RESULTS: MH significantly inhibited HSC activation, as evidenced by the inhibition of cell viability, colony formation and the expression of α-SMA and collagen I. MH caused cell cycle arrest in G2/M phase. Moreover, MH significantly induced apoptosis through regulating the mitochondria-dependent pathway and the activity of caspases. MH treatment significantly increased lysosomes and autophagosomes, and enhanced the formation of autophagic vacuoles and autophagic flux. Interestingly, inhibiting autophagy by 3-MA or RNA interference abolished the ability of MH in inhibiting HSC activation. On the other hand, induction of autophagy promoted MH-induced HSC apoptosis. Further study showed that MH-induced HSC apoptosis and autophagy was mediated by the JNK and PI3K/Akt/mTOR pathways. CONCLUSION: Our results demonstrate that MH-induced HSC apoptosis and autophagy may be one of the important mechanisms for its anti-fibrosis effect.

[Protective effect of isoorientin on alcohol-induced hepatic fibrosis in rats].

OBJECTIVE: To observe the effect and mechanism of isoorientin from Gypsophila elegans on alcohol-induced hepatic fibrosis in rats. METHOD: ninety healthy male Wistar rats were randomly divided into six groups: the normal control group, the model control group, the colchicines group (positive control, 1.0 mg x kg(-1) x d(-1)), the high, middle and low-dose isoorientin groups (20, 50, 100 mg x kg(-1) x d(-1)). The normal control group received normal saline, while other groups received alcohol to cause hepatic fibrosis. After 24-weeks treatment, the alanine aminotransferase (ALT), aspartate aminotransferase (AST), Interleukin 6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), hyaluronic acid (HA), laminin (LN), type III precollagen (PCIII), hydroxyproline (Hyp), Myeloperoxidase (MPO), malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) were assayed according to the manufacturer's instructions, the alpha-SMA and TGF-beta1 were detected by western blotting, and the histopathological changes was observed by H&E staining. RESULT: Isoorientin could improve the liver function by decreasing the activity of ALT, AST, IL-6, TNF-alpha, MDA, MPO, HA, LN, PCIII and Hyp (P < 0.05), increasing the activity of SOD and GSH-Px (P < 0.05), and reducing the expression of alpha-SMA and TGF-beta1 (P < 0.05). In addition, the high and middle-dose isoorientin groups showed more remarkable effect CONCLUSION: Isoorientin from G. elegans can protect hepatic fibrosis induced by alcohol.

Protective effect of isoorientin-2″-O-α-L-arabinopyranosyl isolated from Gypsophila elegans on alcohol induced hepatic fibrosis in rats.

Alcohol abuse is one of the major causes of liver fibrosis, which shows a sharply increasing trend worldwide, yet effective therapeutic options for advanced alcohol fibrosis are limited. Recently we investigated the effect of anti-fibrosis by isoorientin-2″-O-α-L-arabinopyranosyl (IOA) isolated from Gypsophila elegans. During the experiment, the model group received alcohol only, and treatment groups received the corresponding drugs plus alcohol respectively, while the normal control group received an equal volume of saline. Analysis at the end of 24-week experiments showed that IOA could significantly improve the liver function, as indicated by decreasing levels of alanine transaminase, aspartate transaminase, alkaline phosphatase, γ-glutamyltransferase, interleukin-6 and tumor necrosis factor-α. Moreover, IOA could effectively inhibit collagen deposition and reduce the pathological tissue damage. Research on mechanism showed that IOA was able to markedly reduce lipid peroxidation, recruit the anti-oxidative defense system, and induce HSC apoptosis by down-regulating bcl-2 mRNA, as well as inhibit the expression of α-smooth muscle actin and transforming growth factor ß1 proteins. In short, our results showed that IOA is effective in attenuating hepatic injury and fibrosis in the alcohol-induced rat model, which should be developed as a new drug to treat liver fibrosis and even cirrhosis.