中醫傳統方劑－－柴胡疏肝散變方於四氯化碳誘導大鼠肝纖維化動物模式探討抑制肝纖維化之療效

A STUDY OF THE ANTIFIBROTIC EFFECTS OF TRADITIONAL CHINESE HERBS —— MODIFIED CHAI-HU-SHU-GAN POWDER ON CARBON TETRACHLORIDE-INDUCED LIVER FIBROSIS IN RATS

本研究擬以四氯化碳誘導大鼠肝纖維化動物模式，探討中醫傳統方劑－柴胡疏肝散變方抗肝纖維化之作用療效。實驗共分為正常對照組、四氯化碳組、低劑量中藥組（1.26 g/kg ）及高劑量中藥組（6.30 g/kg ），除正常對照組外其他組每週管餵兩次四氯化碳（50 % 1 mL/kg ）連續八週，中藥組同時管餵柴胡疏肝散變方以及四氯化碳組管餵等量一次水每週五次。評估指標包括— 肝功能試驗相關之血液生化值（AST 、ALT ）檢測，組織病理病變評分，肝臟總膠原蛋白含量之定量分析，α-smooth muscle actin 免疫組織化學染色法進行活化態肝臟星狀細胞數量分析，及西方氏墨點偵測法（western blot ）分析α-smooth muscle actin 之蛋白表現量。實驗結果發現餵食高劑量中藥組之AST 、ALT （132 ± 38 U/L; 90 ± 26 U/L ）顯著較四氯化碳組（240 ± 130U/L; 142 ± 32 U/L ）降低（p < 0.05 ），組織病變評分，肝纖維化程度中藥組均較四氯化碳組輕微（p < 0.05 ），肝臟組織中總膠原蛋白之定量分析：高劑量中藥組（22.5 ± 1.5 μg/mg ）肝臟中總膠原蛋白含量明顯低於（p < 0.05 ）四氯化碳組（24.1 ± 1.0 μg/mg ），活化態肝臟星狀細胞數量之表現：高劑量中藥組明顯少於四氯化碳組（p < 0.01 ），且western blot 分析結果：高劑量中藥組之 α-smooth muscle actin 之蛋白表現量明顯低於四氯化碳組（p < 0.001 ），然而低劑量中藥組有幾項評估指標，雖無統計上明顯差異但亦有改善之趨勢。因此從以上結果證實；柴胡疏肝散變方可抑制肝組織中膠原蛋白之合成，以達改善肝纖維化病變程度之療效，其作用機轉推測可能為抑制肝臟中活化態星狀細胞之增生及活化。至於作用機轉進一步的探討以及肝纖維化動物病變程度控制和給藥劑量，為往後研究之重點。The purpose of this study was to investigate the antifibrotic effects of modified Chai-Hu-Shu-Gan (CHSG) powder, comprised of Chinese herbs, on chemical-induced liver fibrosis in adult Sprague- Dawley rats. Liver fibrosis was induced by carbon tetrachloride (50 %, l.0 mL/kg, by gavage) twice a week for 8 weeks. Carbon tetrachloride-induced rats were randomly assigned to three groups: saline, low dose CHSG (1.26 g/kg) and high dose CHSG (6.30 g/kg), each given by gavage 5 times a week for 8 weeks, starting from the onset of carbon tetrachloride administration. Therapeutic effects were assessed by serum enzyme activities (AST and ALT), histopathological score of liver fibrosis, determination of liver collagen content and immunos-taining against smooth muscle cell α-actin to count the number of stellate cells. The content of α-smooth muscle actin was assayed by western blot, result shows that treatment with CHSG (6.30 g/kg) had a more significant reduction (p < 0.05) in the content of AST and ALT (132 ± 38 U/L; 90 ± 26 U/L) than carbon tetrachloride-induced rats receiving saline (240 ± 130 U/L; 142 ± 32 U/L). Liver collagen accumulation was also markedly reduced (p < 0.05) by the CHSG (6.30 g/kg) treatment. Histopathological examination revealed that the CHSG (6.30 g/kg) treatment significantly reduced the degree of liver fibrosis. In addition, it is found that the number of α-smooth muscle actin positive stellate cells and the content of α-smooth muscle actin were markedly reduced (p < 0.001) by CHSG (6.30 g/kg) treatment. These results show that modified Chai- Hu-Shu-Gan powder has anti-fibrotic effects on a carbon tetrachloride-induced liver fibrosi s rat model. Therefore, anti-fibrotic mechanisms might inhibit stellate cell proliferation and activation.