柴胡疏肝散抗肝纤维化的实验研究

目的探讨柴胡疏肝散对肝纤维化大鼠模型α-SMA、TGF-β1的影响及其抗纤维化的机制。方法采用40?l4皮下注射，制备肝纤维化模型并以柴胡疏肝散干预，测定肝功能、血清透明质酸（HA）、层黏连蛋白（LN）及肝组织羟脯氨酸（HYP），光镜观察肝组织病理变化，免疫组织化学法检测肝组织α-SMA、TGF-β1表达，RT-PCR检测TGF-β1 mRNA的表达。结果柴胡疏肝散组较模型组：肝功能明显改善，血清HA及LN显著降低，肝组织HYP含量明显少，肝组织纤维化程度明显改善，肝组织α-SMA及TGF-β1表达减少。结论柴胡疏肝散对CCl4诱导的大鼠肝纤维化有良好的防治作用。     

肝泰胶囊抗肝纤维化作用的实验研究

目的 探讨肝泰胶囊对CCl4诱导的肝纤维化大鼠的作用及其抗肝纤维化的可能机制。方法将40只Wistar大鼠随机分为正常对照组、肝纤维化模型组、肝泰组及肝复乐组。采用光镜、电镜观察肝组织病理学改变，VonGieson（VG）染色及胶原面积半定量分析反映各组肝纤维化程度。采用放射免疫法测定血清层粘连蛋白（LN）、Ⅲ型前胶原（PC-Ⅲ）、Ⅳ型前胶原（Ⅳ-C）、透明质酸（HA）的含量，同时用高效液相色谱-电化学（HPLC—ECD）法检测肝组织中去甲肾上腺素（Norepinephrine，NE）的水平。结果肝泰组较模型组比：①肝组织肝纤维化程度明显改善；②肝组织胶原纤维面积明显减少；③血清透明质酸和层粘连蛋白显著降低；④肝功能损伤明显减轻；⑤组织中NE的含量明显高于模型组。结论肝泰对慢性肝损伤有一定的保护作用。     

牛磺酸对四氯化碳诱导大鼠肝纤维化的抑制作用

目的研究牛磺酸的体内抗肝纤维化作用。方法用四氯化碳（ＣＣｌ４）复制大鼠肝纤维化模型;造模开始或造模６周后给予牛磺酸;实验结束后分别测定肝功能、纤维化指标（透明质酸和层粘连素）、肝羟脯氨酸及Ⅰ、Ⅲ型前胶原ｍＲＮＡ含量,并作肝病理检查。结果牛磺酸可显著减轻ＣＣｌ４肝纤维化程度,能明显降低肝羟脯氨酸和Ⅰ、Ⅲ型前胶原ｍＲＮＡ含量,降低血清透明质酸和层粘连素水平,改善肝功能,组织学检查亦显示具有抗肝纤维化作用。结论牛磺酸在体内具有抗肝纤维化的作用,可望用于肝纤维化的防治。     

甲珠对肝纤维化鼠基质金属蛋白酶表达的影响

目的研究甲珠对肝纤维化大鼠基质金属蛋白酶(MMP)-13、基质金属蛋白酶抑制因子(TIMP)-1影响并探讨其抗纤维化机制。方法采用40%CCl4皮下注射,制备肝纤维化模型,并以甲珠分别采用高剂量、中剂量、低剂量进行干预,采用秋水仙碱做对照。测定肝功能、血清透明质酸、层黏连蛋白及肝组织羟脯氨酸,并通过光镜观察肝组织病理变化,免疫组织化学法检测肝组织MMP-13、TIMP-1表达,RT-PCR检测TIMP-1mRNA的表达。结果甲珠各组较模型组:肝功能明显改善:谷氨酸转氨酶及碱性磷酸酶显著降低,总蛋白及白蛋白显著升高;血清透明质酸及层黏连蛋白显著降低(P<0.05);肝组织羟脯氨酸含量明显少(P<0.05);肝组织纤维化程度明显改善(P<0.01);肝组织MMP-13表达增多、TIMP-1表达减少(P<0.05);肝组织TIMP-1-mRNA表达减少(P<0.01)。结论甲珠对CCL4诱导的大鼠肝纤维化有良好的防治作用。     

中药肝长春保肝抗肝纤维化作用的临床研究

观察中药肝长春保肝抗肝纤维化的治疗效果。用肝长春治疗肝硬化患者55例，另设53例用复方丹参治疗为对照组，治疗前后观察临床表现，检测肝功能，测定血清透明质酸（HA），层粘连蛋白（LN），Ⅳ型胶原（Ⅳ-C）。治疗组临床表现改善明显优于对照组（P＜0．05），总有效率优于对照组（P＜0．05）。肝功能改善及HA，LN，Ⅳ-C的降低明显优于对照组。中药肝长春具有明显的保肝，抗肝纤维化作用。     

抗纤Ⅰ号胶囊抗肝纤维化的血清学和形态学实验研究

目的 观察抗纤Ⅰ号胶囊对实验性肝纤维化大鼠的作用及作用机理。方法 用四氯化碳制作大鼠肝纤维化模型，观察血清层粘连蛋白（LN），Ⅲ型前胶原（PCⅢ）、Ⅳ型胶原（Ⅳ-C），透明质酸（HA）、肝功能和肝组织病理变化。结果 治疗组LN、PCⅢ，Ⅳ-C，HA明显降低，白蛋白明显升高，与对照组比较有显著性差异；肝细胞变性，坯 煞费苦心，肝纤维化程度明显轻于对照组。结论 抗纤Ⅰ号胶囊抗肝纤维化作用肯定，其机理之一，可能是通过降低LN、PCⅢ，从而消除肝窦毛细血管化，调节肝脏微循环，达到抗肝损伤，恢复肝功能的作用。     

IFNα-2b对CHB患者血清肝纤维化和肝功能指标的影响

目的 观察IFNα-2b对慢性乙型肝炎（CHB）患者肝纤维化水平和肝功能的影响.方法 对63例CHB患者予IFNα-2b 500 U肌注,隔日1次,连续治疗3个月.治疗前后检测血清肝纤维化指标Ⅲ型前胶原氨基末端肽（PⅢP）、Ⅳ型胶原（CⅣ）、层粘连蛋白（LN）、透明质酸（HA）和肝功能指标ALT、AST、总胆红素（TB）、白蛋白（A）和球蛋白（G）.结果 治疗后CHB患者血清PⅢP、CⅣ、LN、HA及ALT、AST、TB、A水平均显著低于治疗前（P均＜0.01）,血清G水平显著高于治疗前（P＜0.01）.结论 IFNα-2b可改善CHB患者的肝纤维化水平和肝功能.     

γ-干扰素抗肝纤维化实验研究

目的观察重组人γ－干扰素（IFN－γ）治疗大鼠肝纤维化疗效。右法用二甲基亚硝胺（DMN）腹腔注射诱导大鼠肝纤维化模型,染毒开始及染毒3周后分别用IFN－γ须防和治疗肝纤维化,并与正常对照组和染毒对照组相比较,观察各组血清透明质酸（HA）、肝组织羟脯氨酸（Hyp）、病理肝纤维化程度分期及α-平滑肌肌动蛋白（α－SMA）变化。结果IFN-γ预防和治疗组肝纤维化程度明显轻于染毒对照组,α－SMA阳性细胞表达较染毒对照组明显减少,肝Hyp、血清HA水平较染毒对照组亦有显著降低。IFN－γ台疗组和IFN-γ预防组之间各项指标无显著差异。结论IFN-γ对DMN所致大鼠肝纤维化有较好疗效。     

维生素E对肝纤维化大鼠金属蛋白酶组织抑制因子-2、Ⅲ型胶原及纤维连接蛋白表达的影响

目的 研究维生素E（VE）防治肝纤维化作用及可能存在的机制。方法 以四氯化碳制备大鼠肝纤维化模型,用VE乳剂（100mg/kg,2次／周）静脉注射,连续10周,免疫组织化学检测肝组织Ⅲ型胶原、纤维连接蛋白（FN）沉积;原位杂交检测肝内金属蛋白酶组织抑制因子－2（TIMP－2）及α1（Ⅲ）前胶原mRNA表达。结果 治疗组大鼠肝内Ⅲ型胶原蛋白及FN形成较对照组减少;α1（Ⅲ）前胶原及TIMP－2mRNA表达降低,经图像分析,与对照组差异有显著性（P＜0.05）。结论 VE治疗可下调肝纤维化大鼠TIMP02基因表达,减少细胞外基质沉积,有助于实验性肝纤维化的恢复。     

大黄素对肝纤维化大鼠肝功能的影响

目的 研究大黄素对肝纤维化大鼠肝功能的影响。方法 采用40％四氯化碳(CCl_4)给大鼠皮下注射制备肝纤维化模型并以小、中和大剂量大黄素(20、40和80mg/kg体重)治疗,通过常规方法测定大鼠血清谷丙转氨酶(ALT)、碱性磷酸酶(AKP)、总蛋白(TP)、白蛋白(ALB)和球蛋白(G)。结果 与正常组比较,大黄素组大鼠血清ALT、AKP显著降低(P<0.05～0.01);TP、ALB显著升高(P<0.05～0.01);G降低,白蛋白/球蛋白(A/G)比值升高,但无统计学差异。结论 大黄素对肝纤维化大鼠肝功能有一定的保护作用。     

Effects of PPARg agonist pioglitazone on rat hepatic fibrosis

AIM: To investigate effects of pioglitazone on rat hepatic fibrosis and to explore its mechanism. METHODS: Rat hepatic fibrosis was induced by carbontet. achloride (CCI4). Forty Sprague-Dawley rats were divided randomly into 4 groups: control, model, and two treatment (PⅠ, PⅡ) groups. Except for rats in control group, all rats were given subcutaneous injection of 400 mL/L CCI4, twice a wk for 8 wk. Rats in PⅠ and PⅡ groups were also treated with pioglitazone of 3 mg/kg, daily via gastrogavage beginning on the 1^st day and at the end of the 2^nd week, administration of CCI4 respectively. Liver functions (ALT, AST), serum fibrotic markers (HA, LN, PCIII) and hepatic hydroxyproline (HP) concentration were determined respectively. Histochemical staining of formalin-fixed liver sections with HE, Masson-Trichrome, and immunohistochemical staining for m-smooth muscle actin (α-SMA) were performed. Modified Knodell and Chevallier semi-quantitative scoring system (SSS) was used to evaluate necroinflammatory activity and fibrosis degree. RESULTS: Compared with model group, pioglitazone significantly reduced the serum levels of ALT, AST, HA, LN and PCⅢ (P&lt;0.05 or &lt;0.01). The HP concentrations in PⅠ(210.90&#177;24.07 μg/g), and PⅡ (257.36&#177;30.55 μg/g) groups were also lower than those in model group (317.80&#177;36.44) μg/g) (P&lt;0.01). Histologic examination showed that PⅠ and PⅡ groups had milder hepatocellular degeneration, necrosis and infiltration of inflammatory cells, and thinner or less fibrotic septa than did model group. The scores for necroinflammation in P (2.80&#177;1.03), and PⅡ (3.00&#177;1.05) groups were significantly reduced as compared with model group (4.88&#177;2.30) (P&lt;0.05 or &lt;0.01); the fibrosis scores in PⅠ (3.40&#177;1.65), and PⅡ (4.60&#177;1.35) groups were also markedly lower than those in model group (7.00&#177;3.21) (P&lt;0.05 or &lt;0.01). Immunohistochemical staining showed that expression of α-SMA in PⅠ and PⅡ groups was ameliorated dramatically compared with model group. CONCLUSION: PPARγ, agonist pioglitazone greatly retards the progression of rat hepatic fibrosis induced by CCI4 through inhibition of HSC activation and amelioration of hepatocyte necroinflammation in rats.     

Morphological and serum hyaluronic acid, laminin and type IV collagen changes in dimethylnitrosamine-induced hepatic fibrosis of rats

AIM: To study the morphological and serum hyaluronic acid (HA), laminin (LN), and type IV collagen changes in hepatic fibrosis of rats induced by dimethylnitrosamine (DMN). METHODS: The rat model of liver fibrosis was induced by DMN. Serum HA, type IV collagen, and LN were measured by ELISA. The liver/weight index and morphological changes were examined under electron microscope on d 7, 14, 21, and 28 by immunohistochemical alpha smooth muscle actin alpha-SMA staining as well as Sirius-red and HE staining. RESULTS: The levels of serum HA, type IV collagen and LN significantly increased from d 7 to d 28 (P = 0.043). The liver/weight index increased on d 7 and decreased on d 28. In the model group, the rat liver stained with HE and Sirius-red showed evident hemorrhage and necrosis in the central vein of hepatic 10 lobules on d 7. Thin fibrotic septa were formed joining central areas of the liver on d 14. The number of alpha-SMA positive cells was markedly increased in the model group. Transitional hepatic stellate cells were observed under electron microscope. All rats in the model group showed micronodular fibrosis in the hepatic parenchyma and a network of alpha-SMA positive cells. Typical myofibroblasts were embedded in the core of a fibrous septum. Compared to the control group, the area-density percentage of collagen fibrosis and pathologic grading were significantly different in the model group (P<0.05) on different d (7, 14, and 28). The area-density percentage of collagen fibrosis in hepatic tissue had a positive correlation with the levels of serum HA, LN, and type IV collagen. CONCLUSION: The morphological and serum HA, type IV collagen, and LN are changed in DMN-induced liver fibrosis in rats.     

Cystamine ameliorates liver fibrosis induced by carbon tetrachloride via inhibition of tissue transglutaminase

AIM To investigate the anti-fibrosis effect of the tissue transglutaminase (tTG) specific inhibitor cystamine on liver fibrosis.METHODS Sixty-eight male Sprague Dawley rats were divided into three groups normal control, liver fibrosis control and cystamine-treated group. Liver fibrosis was induced by intraperitoneal injection of carbon tetrachloride (CCl4), and Cystamine was administrated by intraperitoneal injection starting 2 d before the first administration of CCl4. Animals in each group were further divided into 2 subgroups according to two time points of 4 wk and 8 wk after treatment. Hepatic function, pathological evaluation (semi-quantitative scoring system, SSS) and liver hydroxyproline (Hyp)content were examined. Real-time PCR was used to detect the expression of tTG, smooth muscle alpha actin (α-SMA), tissue inhibitor of metalloproteinase 1 (TIMP-1)and collagen-1 mRNA. The expressions of tTG and α-SMA protein were detected by Western Blotting.RESULTS Eight weeks after treatment, the SSS score of liver was significantly less in the cystamine group than that in the fibrosis control group (P ＜ 0.01). The levels of alanine aminotransferase (ALT) and total bile acid (TBA)at the 4 wk and 8 wk time points were decreased in the cystamine group compared with those in fibrosis controls (P ＜ 0.01). Liver hydroxyproline content at the 4 wk and 8 wk time points showed a substantial reduction in the cystamine group compared to fibrosis controls (P ＜ 0.01).The expression of tTG, α-SMA, collagen-1, TIMP-1 mRNA and tTG, as well as α-SMA protein was downregulated in the cystamine group compared to fibrosis controls.CONCLUSION Cystamine can ameliorate CCl4 induced liver fibrosis and protect hepatic function. The possible mechanism is related to the reduced synthesis of the extracellular matrix (ECM) caused by the inhibition of hepatic stellate cell activation and decreased expression of TIMP-1.     

肝窦毛细血管化在二甲基亚硝胺大鼠肝纤维化门静脉高压形成中的作用

目的 研究肝窦毛细血管化在二甲基亚硝胺(DMN)大鼠肝纤维化门静脉高压形成中的作用。方法经4周12次腹腔注射DMN制备火鼠肝纤维化模型,应用电镜技术、免疫组织化学方法结合大鼠门静脉压力测定,24周动态分析肝纤维化形成过程中肝窦毛细血管化与门静脉压力变化的相关性。结果 大鼠门静脉压力随着造模的进行不断升高,造模4周时达(1.10 ±0.18)kPa,明显高于对照组(0.52±0.04)kPa(t=6.41.P<0.0 1)。造模停止后,大鼠门静脉压力逐渐恢复正常。其动态变化与电镜下肝窦毛细血管化的变化规律一致;与反映肝窦内皮表型改变的第Ⅷ因子相关抗原的动态变化成正相关(r=0.833,P<0.01);与反映肝窦内皮下基底膜形成的层黏连蛋白的动态变化成正相关(r=0.953,P<0.01);与反映肝窦壁星状细胞活化收缩的α-平滑肌肌动蛋白的动态变化成正相关(r=0.919,P<0.01)。结论 肝窦毛细血管化是DMN肝纤维化模型门静脉高压产生的主要原因。     

Estrogen reduces CCL4- induced liver fibrosis in rats

AIM：Chronic liver diseasis,such as fibrosis or cirrhosis,are more common in men than in women,This gender difference may be related to the effects of sex hormones on the liver.The aim of the present work was to investigate the effects of estrogen on CCL4-induced fibrosis of the liver in rats.METHODS:Liver fibrosis was induced in male,female and ovariectomized rats by CCL4 administration.All the groups were treated with estradiol(1mg/kg)twice weekly.And tamoxifen was given to male fibrosis model.At the end of 8weeks,all the rats were killed to study serum indicators and the livers.RESULTS:Estradiol treatment reduced aspartate aminotransferase(AST),alanine aminotransferase(ALT),hyaluronic acid(HA)and typeIVcollagen(CIV)in sera,suppressed hepatic collagen content,decreased the areas of hepatic stellate cells(HSC)positive forα-smooth muscle actin(α-SMA).and lowered the synthesis of hepatic typeⅠcollagen significantly in both sexes and ovariectomy fibrotic rats induced byCCL4administration.Whereas.tamoxifen had the opposite effect.The fibrotic response of the female liver to CCL4 treatment was significantly weaker than that of male liver.CONCLUSION:Estadiol reducese CCL4-induced hepatic fibrosis in rats.The antifibrogenic role of estrogen in the liver may be one reason for the sex associated differences in the progression from hepatic fibrosis to cirrhosis.     

Effects of Guiyuanfang and autologous transplantation of bone marrow stem cells on rats with liver fibrosis

AIM: To investigate the therapeutic effects of Guiyuanfang and bone marrow stem cells (BMSCs) on rats with liver fibrosis. METHODS: Liver fibrosis model was induced by carbon tetrachloride, ethanol, high lipid and assessed biochemically and histologically. Liver function and hydroxyproline contents of liver tissue were determined. Serum hyaluronic acid (HA) level and procollagen Ⅲ level were performed by radioimmunoassay. The VG staining was used to evaluate the collagen deposit in the liver. Immunohistochemical SABC methods were used to detect transplanted BMSCs and expression of urokinase plasminogen activator (uPA). RESULTS: Serum transaminase level and liver fibrosis in rats were markedly reduced by Guiyuanfang and BMSCs. HA level and procollagen Ⅲlevel were also reduced obviously, compared to model rats (HA: 47.18±10.97 ng/mL, 48.96±14.79 ng/mL; PCⅢ: 22.48±5.46 ng/mL, 26.90±3.35 ng/mL; P<0.05). Hydroxyproline contents of liver tissue in both BMSCs group and Guiyuanfang group were far lower than that of model group (1 227.2±43.1 μg/g liver tissue, 1390.8±156.3 μg/g liver tissue; P<0.01). After treatment fibrosis scores were also reduced. Both Guiyuanfang and BMSCs could increase the expression of uPA. The transplanted BMSCs could engraft, survive, and proliferate in the liver. CONCLUSION: Guiyuanfang protects against liver fibrosis. Transplanted BMSCs may engraft, survive, and proliferate in the fibrosis livers indefinitely. Guiyuanfang may synergize with BMSCs to improve recovery from liver fibrosis.     

Effects of extract from Ginkgo biloba on carbon tetrachloride-induced liver injury in rats

AIM: To study the effects of extract from Ginkgo biloba (EGb) containing 22% flavonoid and 5% terpenoid on chronic liver injury and liver fibrosis of rats induced by carbon tetrachloride (CCl4). METHODS: All rats were randomly divided into control group, CCl4-treated group, colchicine-treated group and EGb-protected group. Chronic liver injury was induced in experimental groups by subcutaneous injection of CCl4 and fed with chows premixed with 79.5% corn powder, 20% lard and 0.5% cholesterol (v/v). EGb-protected group was treated with EGb (0.5 g/kg body weight per day) for 7 wk. At the end of wk 8, all the rats were killed. Liver function, liver fibrosis, oxidative stress and expression of transforming growth factorβ1 (TGF-β1), a-smooth muscle actin (α-SMA) and typeⅠcollagens in liver were determined. In addition, pathology changes of liver tissue were observed under light microscope. RESULTS: The levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and albumin (Alb) in EGb-protected group were notably improved as compared with the CCL4-treated group (P < 0.01). The contents of serum hyaluronic acid (HA), typeⅢprocollagen (PCⅢ), typeⅣcollagen (CIV) and the expression of hepatic tissue TGF-β1,α-SMA and typeⅠcollagen in EGb-protected group were significantly lower than those in CCL4-treated groups (P < 0.05, P < 0.01). The degrees of liver fibrosis in EGb-protected groups were lower than those in CCL4-treated groups (6.58±1.25 vs 9.52±2.06, P < 0.05). Compared to the CCL4-treated group, the levels of plasma glutathoine peroxidase (Se-GSH-Px), superoxide dismutase (SOD) and malondialdehyde (MDA) were strikingly improved also in EGb-protected group (P < 0.05, P < 0.01). CONCLUSION: EGb resists oxidative stress and thereby reduces chronic liver injury and liver fibrosis in rats with liver injury induced by CCl4