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Effects of Soothing Liver and Invigorating Spleen Recipes on Expression of the Related Gene and Protein on SREBP-1c Signaling Pathway in Hepatocytes and Kupffer Cells of NAFLD Rats

Author: HuangJin
Tutor: YangQinHe
School: Jinan University
Course: Traditional Chinese Medicine
Keywords: Nonalcoholic fatty liver disease Hepatocytes Kupffer cells SREBP-1c signalpathway Soothing liver and invigorating spleen recipes Chaihu Shugan Powder Shen LingBaizhu Powder Rats
CLC: R285.5
Type: Master's thesis
Year: 2013
Downloads: 22
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Abstract


Objective:To observe effects of the soothing liver and invigorating spleen recipes on expression levelsof the related genes and proteins of the SREBP-1c signal pathways in hepatocytes and Kupffercells of the nonalcoholic fatty liver disease(NAFLD) rats, to explore the pathogenesis of NAFLDand the intervention effects of the soothing liver and invigorating spleen recipes.Methods:After one week adaptive feed,75male SD rats were randomly divided into5groups:normal group, model group, soothing liver group, invigorating spleen group, soothing liver andinvigorating spleen recipes integrated group. The normal group was fed basal diet and othergroups were fed with high-fat diet one time everyday. Meanwhile, intervening drug groupsthrough separarely feeding soothing liver recipes, invigorating spleen recipes and integratedrecipes(10mL/kg BW): soothing liver group(receiving gavage of Chaihu Shugan Powder9.6g/kg), invigorating spleen group(receiving gavage of Shen Ling Baizhu Powder30g/kg),integrated group(receiving gavage of Chaihu Shugan Powder and Shen Ling Baizhu Powdercombination recipes39.6g/kg), the normal group and the model group were gavaged withcorrespondence volume distilled water. At the end of eight weeks,9rats were taken fromrespectively each group were anesthetized by3%sodium pentobarbital, the levels of serumlipids, liver lipids and serum aminotransferase were measured by automatic biochemicalanalyzer; Liver pathology was analyzed by HE and Oil red O staining. Another6rats of eachgroup were taken respectively and collagenase (TypeⅣ) was perfused to digest liver tissue withthe circulation in vitro. Hepatocytes were separated by low-speed centrifugation, and Kuppffercells were separated by methods of differential and density gradient centrifugation, SREBP-1c,SCD-1mRNA was assayed by real-time fluorescent quantitative polymerase chain reaction(RTQ-PCR). SREBP-1c, SCD-1protein was detected by Western blot.Results:1. Liver histopathologyNormal group was no obvious lesions and was a little fat droplets stained by Oil-red O.There is structural integrity of liver tissue, hepatic lobules clear cells arranged in neat cable.Model group show medium fatty degeneration in hepatocytes and it has lots of fat dropletsstained by Oil-red O, which is significantly worse than normal group. Swelling of the liver cellswere round, the treatment groups pathology observed between the normal group and modelgroup. 2. Determination of ALT and AST contentCompared with normal group, there was a increase of serum ALT, AST in model group, butthere was no significant difference (P>0.05). There was aslo no significantly difference betweenthe model group and the intervention groups(P>0.05). The results show that ALT, AST of NAFLDrats were not necessarily increased. This is also reflected from the other side, although the theNAFLD rats existence of serious lipid metabolism disorders, liver cells of a large number of fataccumulation, liver cells has not yet reached a severe necrosis of the extent of the damage.3. Determination of blood lipids and liver fat contentSerum lipids Compared with normal group, the levels of serum TC, TG, LDL-C weresignificantly higher (P<0.01), the level of serum HDL-C was significantly decreased in modelgroup (P<0.01). Group compared with model group, the levels of serum TC, LDL-C weresignificantly decreased in the treatment groups(P<0.05,P<0.01), the level of serum TG wassignificantly decreased(P<0.01) and the level of serum HDL-C was significantly decreased insoothing liver group (P<0.01). Comparison of the treatment groups, the level of serum HDL-C ofsoothing liver group was higher than invigorating spleen group and integrated group(P<0.01),the level of serum LDL-C soothing liver group was lower than invigorating spleen group andintegrated group(P<0.05,P<0.01). The results show that there is a serious disorder of lipid metabolism inNAFLD rats, soothing liver and invigorating spleen recipes can reduced to varying degrees ofSerum lipids in NAFLD rats.Liver lipids Compared with the normal group, the lever of serum TC, TG weresignificantly higher in model group(P<0.01).the levels of serum TC, TG were significantlydecreased in soothing liver group(P<0.05). The results show that the levels of liver lipids weresignificantly increased in NAFLD rats, soothing liver and invigorating spleen recipes canreduced to varying degrees of liver lipids in NAFLD rats.4. The expression levels of genes and proteins related to SREBP-1c signal pathway inhepatocytes and Kupffer cellsCompared with the normal group, SREBP-1c, SCD-1mRNA and protein expressions ofhepatocytes and Kupffer cells were significantly increased in the normal group (P<0.01),Compared with model group, SREBP-1c, SCD-1mRNA and protein expressions of hepatocytesand Kupffer cells were significantly decreased in the treatment groups (P<0.01). Comparison ofthe treatment groups, SREBP-1c, SCD-1mRNA expressions of hepatocytes in soothing livergroup were lower than invigorating spleen group(P<0.05), SREBP-1c mRNA of Kupffer cells insoothing liver group was lower than integrated group(P<0.05), SCD-1mRNA of Kupffer cells insoothing liver group was lower than invigorating spleen group(P<0.05). The results show that theactivation of SREBP-1c signal pathway in hepatocytes and Kupffer cells may be an important mechanism for the formation of NAFLD. SREBP-1c, SCD-1mRNA and protein may beimportant targets for prevention and treatment NAFLD by soothing liver and invigorating spleenrecipes.Conclusion:1. Rat model of NAFLD could be successfully established by high-fat diet intervention for8weeks. This model is similar to common human NAFLD type in the changes on pathogenesis,liver biopsy, serum lipids, liver lipids and serum aminotransferase. It is more appropriateanimal model to study the pathogenesis of human NAFLD.2. High-fat diet is a major cause of common human NAFLD Type, the soothing liver andinvigorating spleen may be important therapeutic methods of NAFLD.3. Soothing liver and Invigorating spleen recipes have good effect on NAFLD, which may ofdue to regulation of lipid metabolism disorders and reducing hepatic lipid deposition.4. Hepatocytes and Kupffer cells signal pathway activation of SREBP-1c, raised TC, TGsynthesis, excessive lipid deposited in the liver may be one of the pathogenesis of NAFLD.SREBP-1c, SCD-1mRNA and protein may be effective targets for potential treatment ofNAFLD.5. Soothing liver and Invigorating spleen recipes have good effect on NAFLD, which may ofdue to inhibiting the activation of SREBP-1c signaling pathway in hepatocytes and Kupffercells, downing TC, TG synthesis, Reducing hepatic lipid deposition. SREBP-1c, SCD-1mRNA and protein may be effective targets for Soothing liver and Invigorating spleenrecipes to treat NAFLD.

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