Rating of CCl(4)-induced rat liver fibrosis by blood serum glycomics

BACKGROUND: Non-invasive staging of human liver fibrosis is a desirable objective that remains under extensive evaluation. Animal model systems are often used for studying human liver disease and screening antifibrotic compounds. The aim of the present study was to investigate the potential use of serum N-glycan profiles to evaluate liver fibrosis in a rat model. METHODS: Liver fibrosis and cirrhosis were induced in rats by oral administration of CCl(4). Liver injury was assessed biochemically (alanine aminotransferase [ALT] activity, aspartate aminotransferase [AST] activity and total bilirubin) and histologically. The N-glycan profile (GlycoTest) was performed using DNA sequencer-assisted-fluorophore-assisted carbohydrate electrophoresis technology. In parallel, the effect of cotreatment with antifibrotic interferon-gamma (IFN-gamma) was studied. RESULTS: The biopsy scoring system showed that CCl(4) induced early fibrosis (F < 1-2) in rats after 3 weeks of treatment, and cirrhosis (F4) after 12 weeks. Significant increases in ALT activity, AST activity and total bilirubin levels were detected only after 12 weeks of CCl(4) treatment. GlycoTest showed three glycans were significantly altered in the CCl(4)-goup. Peak 3 started at week 6, at an early stage in fibrosis development (F < 1-2), whereas peaks 4 and 5 occurred at week 9, at which time mild liver fibrosis (F = 1-2) had developed. The changes in the CCl(4)-IFN-gamma group were intermediate between the CCl(4)- and the control groups. CONCLUSION: The GlycoTest is much more sensitive than biochemical tests for evaluating liver fibrosis/cirrhosis in the rat model. The test can also be used as a non-invasive marker for screening and monitoring the antifibrotic activity of potential therapeutic compounds.     

Ginkgo biloba extract reverses CCl4-induced liver fibrosis in rats

AIM: To study the reversing effect of Ginkgo biloba extract (GbE) on established liver fibrosis in rats. METHODS: Following confirmation of CCI4-induced liver fibrosis, GbE or saline was administrated to the rats for 4 weeks. The remaining rats received neither CCI 4 norGbE as normal control. The four groups were compared in terms of serum enzymes, tissue damage, expression of αSMA and tissue inhibitor-1 of metalloproteinase (TIMP-1) and metalloproteinase-1 (MMP-1). RESULTS: Compared with saline-treated group, liver fibrosis rats treated with GbE had decreased serum total bilirubin (P&lt;0.01) and aminotransferase levels (P&lt;0.01) and increased levels of serum albumin (P&lt;0.01). Microscopic studies revealed that the livers of rats receiving GbE showed allieviation in fibrosis (P&lt;0.05) as well as expression of αSMA (P&lt;0.01). The liver collagen and reticulum contents were lower in rats treated with GbE than saline-treated group (P&lt;0.01). RT-PCR revealed that the level of TIMP-1 decreased while the level of MMP-1 increased in GbE group. CONCLUSION: Administration of GbE improved CCI4-induced liver fibrosis. It is possibly attributed to its effect of inhibiting the expression of TIMP-1 and promoting the apoptosis of hepatic stellate cells.     

疏肝健脾方对四氯化碳诱导肝纤维化大鼠血清代谢组学的影响

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Transplantation of bone marrow cells reduces CCl4-induced liver fibrosis in mice

We investigated the effect of bone marrow cell (BMC) transplantation on established liver fibrosis. BMCs of green fluorescent protein (GFP) mice were transplanted into 4-week carbon tetrachloride (CCl4)-treated C57BL6 mice through the tail vein, and the mice were treated for 4 more weeks with CCl4 (total, 8 weeks). Sirius red and GFP staining clearly indicated migrated BMCs existing along with fibers, with strong expression of matrix metalloproteinase (MMP)-9 shown by anti-MMP-9 antibodies and in situ hybridization. Double fluorescent immunohistochemistry showed the expression of MMP-9 on the GFP-positive cell surface. Film in situ zymographic analysis revealed strong gelatinolytic activity in the periportal area coinciding with the location of MMP-9-positive BMCs. Four weeks after BMC transplantation, mice had significantly reduced liver fibrosis, as assessed by hydroxyproline content of the livers, compared to that of mice treated with CCl4 alone. Subpopulation of Liv8-negative BMCs was responsible for this fibrolytic effect. In conclusion, mice with BMC transplants with continuous CCl4 injection had reduced liver fibrosis and a significantly improved survival rate after BMC transplantation compared with mice treated with CCl4 alone. This finding introduces a new concept for the therapy of liver fibrosis.     

Black bean extract ameliorates liver fibrosis in rats with CCl4-induced injury

We assessed the anti-fibrotic effects of methanolic black bean extract antioxidants in a carbon tetrachloride (CCl4) liver injury model in rats. Experimentally intoxicated animals received CCl4 for eight weeks, the reference and test groups received daily intragastric quercetin or daily intragastric black bean extract. Liver fibrosis was assessed and quantified using morphometric analysis. Expression of fibrosis related genes was measured by real time RT-PCR. Qualitative and quantitative histological analysis showed that administration of 70 mg/kg b.w. of black bean extract reduced hepatic fibrosis index by 18% compared to positive controls (P 0.006), as a result of a decrease in type I (44.3% less, P 0.03) and type IV (68.9% less, P 0.049) collagen gene expression compared to CCl4-injured and Quercetin treated rats. In conclusion, we provide evidence that this methanol black bean extract ameliorates liver fibrosis and types I and IV collagen gene expression, in the animal model used. PRACTICAL APPLICATIONS: The compounds contained in this black bean extract exhibited strong antifibrotic effects in the CCl4 chronic liver injury model used; considering that this compounds are contained in a leguminous that has been used in human diet for a long time, their toxic potential should be very low, and this characteristic should favor their potential use in some other chronic or degenerative states that include an increase in inflammation and oxidative burst in their pathogenesis. Another possible application of this kind of extract could be its use as an antimicrobial or even antiparasitic therapeutic agent, although it is purely speculative.     

依达拉奉对实验性肝纤维化大鼠脂质过氧化的影响

目的探讨依达拉奉（EDA）对实验性肝纤维化大鼠脂质过氧化的影响。方法以四氯化碳诱导大鼠肝纤维化模型。30只Sprague—Dawley大鼠随机分为对照组（10只）、肝纤维化模型组（10只）和EDA防治组（10只）。检测各组大鼠血清谷丙转氨酶、谷草转氨酶水平及肝组织羟脯氨酸、丙二醛含量和超氧化物歧化酶活性。结果EDA防治大鼠血清谷丙转氨酶和谷草转氨酶水平分别为714．2±28．2U／L和766．0±11．0U／L,较模型组（1110．3±45．9U／L和1640．3±26．7U／L,P〈0．05和P〈0．01）明显降低;EDA组大鼠肝组织羟脯氨酸和丙二醛含量分别为0．4±0．1μg／mg和5．5±2．3nmol／gl,较模型组（0．8±0．1μg／mg和7．5±2．1nmol／gl,P均〈0．01）显著下降;EDA组大鼠超氧化物歧化酶活性为129．7±2．3u／g,较模型组（933±3．9u／g,P〈0．01）明显升高;EDA组和模型组大鼠肝组织纤维化评分分别为2．7±1．0和3．5±0．7,差异显著（P〈0．01）。结论EDA对大鼠肝纤维化有一定的防治作用,其机制很可能与抗脂质过氧化损伤有关。     

脾切除对大鼠实验性肝纤维化的影响

目的：研究脾脏在大鼠四氯化碳肝纤维化模型中对肝纤维化形成的影响。方法：Ｗｉｓｔａｒ大鼠２３６只，随机分为模型组，预先脾切除组，注射四氯化碳后６．９、１２周分别行脾切除组及相应的假手术组。应用光镜观察组织学改变，放免法测定肝匀浆Ⅲ型前胶原、透明质酸、层粘连蛋白水平和肝组织羟脯氨酸含量，并结合胶原计算机图像分析来评价脾脏切除对四氯化碳诱发大鼠肝纤维化的影响。结果：脾脏切除能减轻肝细胞损伤。减少细胞外基质成分在肝内的沉积，延缓肝纤维化的发生。脾切除组肝纤维化进展较不切脾模型组延缓１个实验阶段（３周），即使在造模过程肝纤维化尚未完全形成时切除脾脏。对肝纤维化也有一定的减轻和延缓作用．结论：脾脏切除可在一定程度上减轻和延缓大鼠四氯化碳所致肝纤维化。     

肝纤维化大鼠肝组织Smads基因表达状况及意义

目的:研究CCl4诱导的肝纤维化大鼠肝组织Smad基因表达的变化及其意义.方法:80只健康雄性SD大鼠分为2组:正常组(C组,n=40)和模型组(M组,n=40).以CCl4 sc 法诱导肝纤维化,HE和VG胶原染色观察肝脏胶原沉积情况,原位杂交法及免疫组化法检测肝组织Smads分子表达水平变化.结果:与C组比较,M组大鼠肝脏组织学积分显著增加(3.29±0.68 vs0,P<0.05),平均胶原面积显著增加(290.86±89.37 μm2 vs 56.12±21.45 μm2,P<0.01),肝组织Smad4蛋白表达率较C组明显增加(4.27%±0.43% vs 2.86%±0.86%,P<0.05),而Smad7蛋白表达虽然增加,但水平低下;Smad 3 mRNA表达A值明显增加(0.167±0.092 vs 0.010±0.002,P<0.05),Smad4 mRNA表达A值也明显增加(0.24 1±0.098 vs 0.021±0.004,P<0.05),Smad6、Smad7 mRNA虽然增加,但表达水平仍然低下.结论:实验性大鼠肝纤维化存在肝脏Smads分子表达水平的比例失调,TGF-Smad信号通路可能参与了肝纤维化的形成与发展.     

Protective effect of carnosol on CCl(4)-induced acute liver damage in rats

BACKGROUND: We recently reported that (Lamiaceae) may alleviate CCl(4)-induced acute hepatotoxicity in rats, possibly blocking the formation of free radicals generated during CCl(4) metabolism. Carnosol, one of the main constituents of Rosmarinus, has been shown to have antioxidant and scavenging activities. Therefore, it is plausible to expect that carnosol may mediate some of the effects of Rosmarinus on oxidative stress consequences induced by CCl(4) in the liver. DESIGN: We evaluated the effectiveness of carnosol to normalize biochemical and histological parameters of CCl(4)-induced acute liver injury. METHODS: Male Sprague Dawley rats (n = 5) injured by CCl(4) (oral dose 4 g/kg of body weight) were treated with a single intraperitoneal dose (5 mg/kg) of carnosol. Twenty-four hours later, the rats were anaesthetized deeply to obtain the liver and blood, and biochemical and histological parameters of liver injury were evaluated. RESULTS: Carnosol normalized bilirubin plasma levels, reduced malondialdehyde (MDA) content in the liver by 69%, reduced alanine aminotransferase (ALT) activity in plasma by 50%, and partially prevented the fall of liver glycogen content and distortion of the liver parenchyma. CONCLUSIONS: Carnosol prevents acute liver damage, possibly by improving the structural integrity of the hepatocytes. To achieve this, carnosol could scavenge free radicals induced by CCl(4), consequently avoiding the propagation of lipid peroxides. It is suggested that at least some of the beneficial properties of Rosmarinus officinalis are due to carnosol.     

Effects of platelet-rich plasma on liver regeneration in CCl4-induced hepatotoxicity model

Numerous bioactive growth factors and cytokines in platelet-rich plasma (PRP) have recently made it an attractive biomaterial for therapeutic purposes. These growth factors have the potential to regenerate the injured tissues. The aim of this study was to investigate the therapeutic effects of PRP in hepatotoxic animal model. Hepatotoxicity was induced in rats by oral administration of 4 mL/kg/week of CCl₄ diluted 1:1 in corn oil for 10 weeks. To confirm the hepatotoxicity, 24 h after the last CCl₄ administration, blood samples were collected via cardiac puncture to assess the serum levels of alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, albumin, total protein, and total bilirubin. Twenty-four hours after blood collection, the experimental animals received a single injection of PRP (1 mL) via the anterior mesenteric vein. One week later, all biochemical tests were performed again, and the rats were scarified and their livers were removed, prepared histologically, and stained. The stereological analyses were performed to evaluate the effects of PRP on histopathological features of CCl₄-treated livers. The results were compared statistically with the corresponding control and CCl₄+normal saline (NS)-treated animals. A significant decrease in the number and volume of hepatocytes (p = 0.01), and also a reduction in the volume of sinusoids (p = 0.001) and connective tissue (p = 0.04), were observed in the PRP-treated animals compared with the CCl₄+NS-treated ones. Our findings demonstrated that application of PRP had beneficial effects on CCl₄-induced fibrosis; however, it had detrimental effects on the total number of hepatocytes and the volume of hepatocytes and sinusoidal spaces.     

软肝冲剂对肝纤维化大鼠组织金属蛋白酶抑制剂的影响

目的：探讨软肝冲剂治疗肝纤维化和肝硬化的机制是否与抑制TIMP-1的表达有关。方法：用CCl4诱导大鼠的肝纤维化模型（病理分期为0-Ⅴ期），将大鼠随机分成两组，分别用软肝冲剂和生理盐水治疗2个月后。对肝组织TIMP-1mRNA的表达进行分析，并检测羟脯氨酸（Hyp）的量，同时检测血清中TIMP-1和TGF-β1的浓度。结果：软肝冲剂能够降低大鼠肝组织中TIMP-1mRNA的表达和胶原蛋白的含量，同时血清中TIMP-1和TGF-β1较CCl4诱导的模型组和生理盐水治疗组显著降低（P〈0．05）。结论：软肝冲剂治疗肝纤维化可能通过阻止TIMP-1的表达起作用。     

不同浓度CCl4注射联合饮用乙醇诱导大鼠肝硬化模型研究

目的 探索不同浓度CCl4联合饮用乙醇制备大鼠肝硬化模型,寻求最佳的药物和乙醇浓度。方法 取90只SD大鼠, 分为对照组10只和实验组A组、B组、C组和D组,每组20只,给予不同浓度的CCl4油溶液腹腔注射,同时以不同浓度的乙醇溶液为饮用水,制备肝硬化模型。结果 在实验12 w末,A组大鼠死亡10只, B组死亡8只,C组死亡4只,D组死亡5只;实验A组肝细胞走向紊乱,纤维组织增生,纤维间隔形成。有假小叶形成,肝细胞大小不一,呈点灶状坏死,肝细胞凋亡、再生,汇管区内炎细胞浸润;B组与A组变化相似;C组肝细胞走向紊乱,纤维间隔形成,但无假小叶形成,肝细胞脂肪变性,纤维组织增生,肝细胞凋亡,汇管区内炎细胞浸润;D组与C组表现类似。结论 适当浓度的CCl4油溶液结合乙醇溶液为饮用水诱导大鼠肝硬化模型可显著提高造模成功率。     

磁共振扩散加权成像诊断四氯化碳诱导的兔肝纤维化的价值分析

目的探讨磁共振扩散加权成像(DWI)对肝纤维化的诊断价值。方法采用四氯化碳注射法建立家免肝纤维化模型每次实验抽取模型组家免7□8只和对照组家免2□3只进行DWI检查,共对30只肝纤维化成模组和10只对照组家免进行了DWI检查测算不同b值(分别为300、500和1000s/mm~2)时DWI的表观扩散系数(ADC)值;行DWI后12h内处死家免行肝组织病理学检查,按肝纤维化病理分期结果进行分组,比较不同分期动物ADC值的差异,采用Spearman相关分析探讨ADC值变化和纤维化分期之间的相关性,运用受试者工作特性(ROC)曲线评估ADC值预测S_2及以上肝纤维化和S_3及以上肝纤维化的诊断效能。结果ADC值与纤维化分期之间呈负相关性取b值=500 s/mm~2时相关性最高(r=0.795,P=0.000);在不同b值情况下肝纤维化≤S_1与≥S_2之间、纤维化≤S_2与≥S_3之间肝脏ADC值差异均有统计学意义(均P0.05);当b值=500 s/mm~2时,ADC值诊断≥S_2肝纤维化的ROC曲线下面积(AUC)为0.912,以ADC值=1.58×10~(-3)mm~2/s为截断点,其诊断肝纤维化的敏感性为90.2%特异性为75.0%;ADC值诊断≥S_3肝纤维化的AUC为0.920,以ADC值=1.43×10~(-3)mm~2/s为截断点,其敏感性为93.0%特异性为80.0%。结论ADC值可以用于诊断肝纤维化分期值得进一步研究。     

Influence factors of serum fibrosis markers in liver fibrosis

AIM: To analyze the factors which influence the serum levels of hyaluronic acid (HA), type III pro-collagen (PCIII), laminin (LN) and type IV collagen (C-IV) in liver fibrosis. METHODS: The serum specimens from 141 chronic hepatitis patients were assayed for fibrosis indexes including HA, PCIII, LN and C-IV with radioimmunoassay (RIA) and liver function indexes by an automatic biochemistry analyzer. The patientswere then divided into consistent group and inconsistent group. The patients‘‘clinical manifestations were recorded, routine blood pictures were done by a blood counter and analyzer (AC-900). Liver biopsy specimens were examined path-morphologically. The inner diameters of portal vein, splenic vein and thickness of spleen were all measured by ultrasonography. RESULTS: Sixteen patients (14.16%) had serum fibrosis indexes inconsistent with histological stage of their hepatic fibrosis. Their serum fibrosis indexes did not correlate with the stage of hepatic fibrosis (P&gt;0.05), but were positively correlated with the grade of inflammation (72=12.07, P&lt;0.05). At the same time, serum albumin (ALB) and the ratio of albumin and globulin (A/G) were significantly increased (t=3.06, P&lt;0.01), (t=3.70, P&lt;0.01). Serum levels of glutamic-pyruvic transaminase (ALT), glutamic-oxaloacetic transaminase(AST), γ-glutamyl transferase (GGT) and globulin (GLB) were all significantly decreased (t=-2.45, P&lt;0.05), (t=-2.33,P&lt;0.05), (t=2.08, P&lt;0.05), (t=-3.03, P&lt;0.01). Weary degreealso decreased more obviously (χ^2=7.52, P&lt;0.05), but other clinical manifestations, routine blood indexes, serum levels of alkaline phosphatase (AKP), total bilirubin (TBIL), total protein (TP), width of main portal vein, width of splenic vein and thickness of spleen had no significant change (P&gt;0.05). CONCLUSION: Serum fibrosis indexes can be influenced by the grade of inflammation, some liver function indexes and clinical manifestations. Comprehensive analysis is necessary for its proper interpretation.     

β受体阻断剂卡维地洛抗肝纤维化的作用

目的 研究非选择性β受体阻断荆和α-1受体阻滞剂卡维地洛对抗大鼠肝纤维化的作用及机理.方法 40只sD大鼠分为4组.分别为正常对照组、肝纤维化组、单用卡维地洛组、卡维地洛治疗组,分别测定各组的肝功能[丙氨酸氮基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)、白蛋白/球蛋白(A/G)],肝纤维化血清指标(PC4、HA、CN),肝纤维化组织学指标(HE染色,Masson染色).SP免疫组织化学法测定肝组织α平滑肌肌动蛋白(α-SMA)和RT-PCR法测定肝组织转化生长因子β1 mRNA(TGF-β1 mRNA).结果 与造模组相比,卡维地洛改善了肝功能,血清肝纤维化指标和病理学检查指标都有所下降.结论 卡维地洛能够减轻CCl4致大鼠的肝纤维化.     

Coffee prevents CCl4-induced liver cirrhosis in the rat

Purpose  

Previous clinical observations suggested that coffee may have beneficial effects on the liver. In fact, an inverse relationship between coffee consumption and liver cirrhosis has been reported in humans. However, the causative role of coffee has not been established; therefore, the aim of this work was to study the effect of coffee in an experimental model of liver damage.     

酒精性肝纤维化的血清标志物研究进展

酒精性肝纤维化是由于长期过量摄入酒精使肝细胞发生炎症及坏死等改变,导致纤维结缔组织在肝内过度沉积而引发的一系列病理改变。实验室指标有助于助鉴别酒精性肝病,再进一步联合肝纤维化检验方法可诊断酒精性肝纤维化。其中肝纤维化诊断的"金标准"仍然是肝活组织检查,但因为其有创性,临床实践中开展受限。因此具有诊断价值的血清新型分子标志物近来不断被发现,且取得了较大突破;同时以综合多项临床生化指标、影像学为基础的肝纤维化非创伤性诊断预测模型对肝纤维化的诊疗也带来了新的希望。综述了近年来用血清标志物对酒精性肝纤维化非侵入诊断进行预测评估的研究进展。