Effect Of Emodin On Methionine-choline-deficient Diet Induced Mouse Non-alcoholic Steatohepatitis

Non-alcoholic fatty liver disease(N AFLD) is a chronic liver disease with high prevalence clinically, and its ma in characteristic is fat deposit ion in the liver, which is associated wit h the imbalance of fat intake and decomposition. Wit h the development of economy, the lifestyle and diet structure are changed and the NAFLD incidence is increased accordingly. N on-alcoholic steatohepatit is(N ASH) is a gradually developed disease and is a pathologic subtype of N AFLD. S tatist ically, about 30% ~ 40% of N ASH is comb ined wit h liver fibrosis, 10%~15% of patients with N ASH will develop as liver cirrhosis, and about 9~26% of N ASH liver cirrhosis patients will die in 4~10 years. Nevertheless, there is no efficient treatment on it. Thus, prevention of N ASH timely w ill s ignificant ly reduce the inc idence of liver irrhosis and improve the quality of life.Emodin is extracted from certain tradit iona l C hinese medic ine(such as rhubarb, fleece- flower root) and belongs to the anthraquinone materia l. Studies have shown that emodin has extensive pharmacological effects, such as anti- infla mmation, anti-bacteria, induction of diuresis, repression of blood pressure, ant i- tumor and so on. Two animal models are used for studing N ASH. O ne is high- fat diet induced NASH, and another is methionine-choline- deficient diet induced N ASH. O ur previous data showed that emodin significantly improved high- fat diet induced NASH. However, whether emodin exerts a protective role in MC D- induced N ASH has not been reported yet.Objective To investigate the effect of emodin on MCD diet induced NASH.Method Experime nt I:Establishment of MCD diet induced NASH model 1. After an adaptive feeding for one week, 20 C 57BL/6J mice were randomly divided into two groups: methionine- choline- supple mented diet group(MC S) and methionine- choline defic ient diet group(MC D). The mice were fed with MC S or MC D diet respective ly for 4 weeks; 2. Mouse weight was measured weekly; 3. When the mice were sacrificed, body weight and liver weight were recorded, and the liver tissues and serum were collected; 4. Hematoxylin- eosin(HE) staining was performed to observe the pathologic changes of livers; 5. Plas ma alanine aminotransferase(ALT) and aspartate transaminase(AST) leve ls were tested by automatic biochemical analyzer. Experime nt II:The effect of emodin on MCD diet induced NASH 1. After an adaptive feeding for one week, 38 C57BL/6J mice were randomly divided into three groups: MCS diet plus DMSO injection group(MCS), MC D diet plus DMSO injection group(MC D) and MC D diet plus emodin injection group(MC D-emodin); 2. The mice were fed wit h MC S or MC D diet for 10 days, then peritoneally injected with DMSO or emodin(40 mg/kg) for 18 days consecutive ly; 3. Mouse weight was measured weekly; 4. When the mice were sacrificed, body weight, liver weight and spleen weight were recorded, and the liver and spleen tissues andserum were collected; 5. Hematoxylin- eosin(HE) staining was performed to observe pathologic changes of livers; 6. Plasma ALT, AST, triglyceride(TG), serum total cholesterol(TC), high density lipoprotein(HDL), low dens ity lipoprotein(LDL), alkaline phosphatase(ALP), glucose(GLU) levels were measured by automat ic biochemical analyzer; 7. Interleukin 1β(IL-1β), Interleukin 6(IL-6) and tumor necrosis factor- α(TNF-α) m RNA levels were determined by quant itative real- time PCR.Results Experiment I: 4 weeks after MC D feeding, mice were apathetic. MC D diet fed mice exhib ited a significantly decreased body weight when compared to MCS group(p < 0.05). HE staining showed that there were more lip id accumulation and leukocyte infiltration in livers of MC D diet fed mice relat ive to MCS group. The plasma AST and ALT leve ls in MC D diet fed mice were remarkably increased compared with MC S group(p < 0.05). Experiment II: 1. There was no significant difference in body weight, liver weight and spleen weight between MC D and MC D- emodin group. 2. HE staining showed that there were more lipid accumulation and leukocyte infiltrat ion in livers of MC D diet fed mice compared to MC S group. However, above phenomena were more severe upon emodin injection. 3. Moreover, compared to MC D group, emodin injection remarkably enhanced plasma AST and ALT leve ls and greatly increased IL-1β and IL-6 m RNA levels.Conclusion: Our current study successfully established MC D diet induced N ASH mouse model, as demonstrated by increased lipid accumulat ion and leukocyte infiltration, and impaired liver funct ion. Emodin intervention dramatically aggravated steatosis,increased serum ALT and AST leve ls, and enhanced inflammatory factor expression. Our data indicated that emodin worsened MCD diet induced mouse NASH.