Protective effect of bicyclol against bile duct ligation-induced hepatic fibrosis in rats

AIM: To evaluate the protective effect of bicyclol against bile duct ligation(BDL)-induced hepatic fibrosis in rats.METHODS: Sprague-Dawley male rats underwent BDL and sham-operated animals were used as healthy controls. The BDL rats were divided into two groups which received sterilized PBS or bicyclol(100 mg/kg per day) orally for two consecutive weeks. Serum, urine and bile were collected for biochemical determinations. Liver tissues were collected for histological analysis and a whole genome oligonucleotide microarray assay. Reverse transcription-polymerase chain reaction and Western blotting were used to verify the expression of liver fibrosis-related genes.RESULTS: Treatment with bicyclol significantly reduced liver fibrosis and bile duct proliferation after BDL. The levels of alanine aminotransferase(127.7 ± 72.3 vs 230.4 ± 69.6, P < 0.05) and aspartate amino-transferase(696.8 ± 232.6 vs 1032.6 ± 165.8, P < 0.05) were also decreased by treatment with bicyclol in comparison to PBS. The expression changes of 45 fibrogenic genes and several fibrogenesis-related pathways were reversed by bicyclol in the microarray assay. Bicyclol significantly reduced liver m RNA and/or protein expression levels of collagen 1a1, matrix metalloproteinase 2, tumor necrosis factor, tissue inhibitors of metalloproteinases 2, transforming growth factor-b1 and α-smooth muscle actin.CONCLUSION: Bicyclol significantly attenuates BDLinduced liver fibrosis by reversing fibrogenic gene expression. These findings suggest that bicyclol might be an effective anti-fibrotic drug for the treatment of cholestatic liver disease.

Protective effects of a polymethoxy flavonoids-rich Citrus aurantium peel extract on liver fibrosis induced by bile duct ligation in mice

Objective: To evaluate the possible protective effect of Citrus aurantium peel extract(CAE) against apoptosis in cholestatic liver fibrosis induced by bile duct ligation in mice. Methods: Male ICR mice were divided to 5 groups: 1) Control group(Sham-operated mice), 2) Cholestatic liver injury group induced by bile duct ligation(BDL), 3) BDL mice treated with silymarin(200 mg/kg) for 4 weeks, 4) BDL mice treated with 50 mg/kg CAE for 4 weeks, 5) BDL mice treated with 200 mg/kg CAE for 4 weeks. Mice were sacrificed and liver fibrosis was evaluated by serum and hepatic tissue biochemistry tests and liver histopathological examination. Effects of CAE on inflammation and apoptosis gene regulation were investigated through real-time PCR. CAE effect on lipid metabolism related signaling was determined by western blot analysis. Results: In BDL mice, administration of CAE for 4 weeks markedly attenuated liver fibrosis based on histopathological alteration. Serum and hepatic tissue biochemistry results revealed that CAE(50 and 200 mg/kg) decreased the levels of alanine transaminase, aspartate transaminase, gamma glutamyl transferase, total bilirubin, nitric oxide, and thiobarbituric acid reactive substances. Real-time PCR and western blot analysis showed that CAE regulated inflammation, apoptosis, and lipid metabolism factors increased by BDL. Interleukin family, tumor necrosis factor α, and related apoptosis factors m RNA levels were increased by BDL treatment. However, these increases were suppressed by CAE administration. In addition, CAE effectively increased phosphorylation of AMPactivated protein kinase, nuclear factor E2-related factor 2, and related cytoprotective proteins. Conclusions: CAE can efficiently regulate BDL-induced liver injury with antioxidant, antiinflammatory, and antiapoptotic activities.

Multiparameter magnetic resonance imaging of liver fibrosis in a bile duct ligation mouse model

BACKGROUND Bile duct ligation(BDL)in animals is a classical method for mimicking cholestatic fibrosis.Although different surgical techniques have been described in rats and rabbits,mouse models can be more cost-effective and reproducible for investigating cholestatic fibrosis.Magnetic resonance imaging(MRI)has made great advances for noninvasive assessment of liver fibrosis.More comprehensive liver fibrotic features of BDL on MRI are important.However,the utility of multiparameter MRI to detect liver fibrosis in a BDL mouse model has not been assessed.AIM To evaluate the correlation between the pathological changes and multiparameter MRI characteristics of liver fibrosis in a BDL mouse model.METHODS Twenty-eight healthy adult male balb/c mice were randomly divided into four groups:sham,week 2 BDL,week 4 BDL,and week 6 BDL.Multiparameter MRI sequences,included magnetic resonance cholangiopancreatography,T1-weighted,T2-weighted,T2 mapping,and pre-and post-enhanced T1 mapping,were performed after sham and BDL surgery.Peripheral blood and liver tissue were collected after MRI.For statistical analysis,Student’s t-test and Pearson’s correlation coefficient were used.RESULTS Four mice died after BDL surgery;seven,six,five and six mice were included separately from the four groups.Signal intensities of liver parenchyma showed no difference on TI-and T2-weighted images.Bile duct volume,ΔT1 value,T2 value,and the rate of liver fibrosis increased steadily in week 2 BDL,week 4 BDL and week 6 BDL groups compared with those in the sham group(P<0.01).Alanine aminotransferase and aspartate transaminase levels initially surged after surgery,followed by a gradual decline over time.Strong correlations were found between bile duct volume(r=0.84),T2 value(r=0.78),ΔT1 value(r=0.62),and hepatic fibrosis rate(all P<0.01)in the BDL groups.CONCLUSION The BDL mouse model induces changes that can be observed on MRI.The MRI parameters correlate with the hepatic fibrosis rate and allow for detection of cholestatic fibrosis.

Nrf2 and Snail-1 in the prevention of experimental liver fibrosis by caffeine

AIM:To determine the molecular mechanisms involved in experimental hepatic fibrosis prevention by caffeine(CFA).METHODS:Liver fibrosis was induced in Wistar rats by intraperitoneal thioacetamide or bile duct ligation and they were concomitantly treated with CFA(15 mg/kg per day).Fibrosis and inflammatory cell infiltrate were evaluated and classified by Knodell index.Inflammatory infiltrate was quantified by immunohistochemistry(anti-CD11b).Gene expression was analyzed by quantitative reverse transcription-polymerase chain reaction for collagenⅠ?(Col-1),connective tissue growth factor(CTGF),transforming growth factorβ1(TGF-β1),tumor necrosis factor alpha(TNF-α),interleukin-1(IL-1),IL-6,superoxide dismutase(SOD)and catalase(CAT).Activation of Nrf2 and Snail-1 was analyzed by Westernblot.TNF-αexpression was proved by enzyme-linked immunosorbant assay,CAT activity was performed by zymography.RESULTS:CFA treatment diminished fibrosis index in treated animals.The Knodell index showed both lower fibrosis and necroinflammation.Expression of profibrogenic genes CTGF,Col-1 and TGF-β1 and proinflammatory genes TNF-α,IL-6 and IL-1 was substantially diminished with CFA treatment with less CD11b positive areas.Significantly lower values of transcriptional factor Snail-1 were detected in CFA treated rats compared with cirrhotic rats without treatment;in contrast Nrf2was increased in the presence of CFA.Expression of SOD and CAT was greater in animals treated with CFA showing a strong correlation between mRNA expression and enzyme activity.CONCLUSION:Our results suggest that CFA inhibits the transcriptional factor Snail-1,down-regulating profibrogenic genes,and activates Nrf2 inducing antioxidant enzymes system,preventing inflammation and fibrosis.

Effects of Yinchenhao Decoction on Self-regulation of Renin-angiotensin System by Targeting Angiotensin Converting Enzyme 2 in Bile Duct-ligated Rat Liver

Summary： In order to investigate whether Yinchenhao decoction （YCHD） attenuates hepatic fibro- genesis in the bile duct ligation （BDL） model via recovering and restoring the self-regulation and bal- ance of the renin-angiotensin system （RAS）, 33 specific-pathogen-free （SPF） male Sprague-Dawley rats with common BDL and scission were randomly divided into five groups as follows： G1, the sham group （n=4）; G2, BDL 7-day group （n=5）; G3, BDL＋YCHD 430 mg/mL （n=8）; G4, BDL＋losartan 0.65 mg/mL （ARB group, n=8）; G5, model group （BDL without any treatment, n=8）. YCHD and losartan （10 mL.kgl.day-1） were given by gastric gavage for 16 days following BDL in G3 and G4 groups, respec- tively. The effect of YCHD on liver fibrosis and the detailed molecular mechanisms were assessed by liver function including total bilirubin （TBIL）, direct bilirubin （DBIL）, indirect bilirubin （IDBIL）, alanine aminotransferase （ALT）, and aspartate aminotransferase （AST）. Histological changes were ob-. served by transmission electron microscopy （TEM） and Masson trichrome staining. Western blotting was used to detect the protein expression level of the renin-angiotensin system （RAS） components in- cluding angiotensin converting enzyme （ACE）, angiotensin II type 1 receptor （AT1R）, ACE2, angio- tensin II （Ang II） as well as transforming growth factor 131 （TGF131）. The experimental data were ana- lyzed by principle component analytical method of pattern recognition. The results showed that bio- chemically, serum TBIL, DBIL, IDBIL, ALT and AST levels were markedly increased following BDL as compared with the sham group （P〈0.05）. Serum TBIL, IDBIL and DBIL levels in G3 group were dramatically decreased as compared with G5 and G4 groups （P〈0.05）. Serum AST level in G3 was sig- nificantly lowered than in G5 group （P〈0.05）, but there was no significant difference in ALT among G3, G4 and G5 groups （P〉0.05）. Histologically, livers in G3 group showed less hepatocytes necrosis, less bile duct hyperplasia and less collagen formation than in G4 and G5 groups. The protein expression lev- els of ACE2, ACE, Ang II, AT1R and TGF131 in G2, G3 and G4 groups were significantly higher than in sham group （P〈0.05）, and lower than in G5 group （P〈0.05）. However, the differences among G2, G3 and G4 groups were not significant （P〉0.05）. ACE2 protein expression in G3 group was significantly higher than in G2 group （P〈0.05） and there was no significant difference in comparison with G4 group （P〉0.05）. Moreover, the protein expression of TGF131 in G3 group was significantly lower than in G5 and G4 groups （P〈0.05）. Our findings suggest that the antifibrotic effects of YCHD may be associated with the decreased classical RAS pathway components and TGFI31 downexpression so as to recover and rebuild self-regulation of the RAS by elevating the protein expression of ACE2.

Heme oxygenase-1 overexpression increases liver injury after bile duct ligation in rats

AIM: To investigate the effects of heme oxygenase-1 (HO-1) against oxidant-induced injury caused by bile duct ligation (BDL). METHODS: Either cobalt protoporphyrin (CoPP), a HO-1 inducer, or saline were injected intraperitoneally in male SD-rats. Three days later, BDL or sham-operations were performed. Rats were sacrificed 3 wk after BDL and livers were harvested for histology. Fibrosis was evaluated by sirius red staining and image analysis. Alpha-smooth muscular actin, which indicates activation of stellate cells, was detected by immunohistochemical staining, and cytokine and collagen-Ⅰα (Col-Ⅰα) mRNA expression was detected using RNase protection assays. RESULTS: Serum alanine transaminase increased 8-fold above normal levels one day after BDL. Surprisingly, enzyme release was not reduced in rats receiving CoPP. Liver fibrosis was evaluated 3 wk after BDL and the sirius red-positive area was found to be increased to about 7.8%. However, in CoPP pretreated rats sirius red- positive areas were increased to about 11.7% after BDL. Collagen-Ⅰα and TGF-β mRNA increased significantly by BDL. Again, this effect was increased by HO-1 overexpression.CONCLUSION: Hepatic fibrosis due to BDL is not reduced by the HO-1 inducer CoPP. In contrast, HO-1 overexpression increases liver injury in rats under conditions of experimental chronic cholestasis.

Antioxidant Effect of Sepia Ink Extract on Extrahepatic Cholestasis Induced by Bile Duct Ligation in Rats

Objective The aim of our study was to assess the complications of hepatic fibrosis associated with bile duct ligation and the potential curative role of sepia ink extract in hepatic damage induced by bile duct ligation. Methods Rattus norvegicus rats were divided into 3 groups: Sham-operated group, model rats that underwent common bile duct ligation (BDL), and BDL rats treated orally with sepia ink extract (200 mg/kg body weight) for 7, 14, and 28 d after BDL. Results There was a significant reduction in hepatic enzymes, ALP, GGT, bilirubin levels, and oxidative stress in the BDL group after treatment with sepia ink extract. Collagen deposition reduced after sepia ink extract treatment as compared to BDL groups, suggesting that the liver was repaired. Histopathological examination of liver treated with sepia ink extract showed moderate degeneration in the hepatic architecture and mild degeneration in hepatocytes as compared to BDL groups. Conclusion Sepia ink extract provides a curative effect and an antioxidant capacity on BDL rats and could ameliorate the complications of liver cholestasis.

Comparison of murine cirrhosis models induced by hepatotoxin administration and common bile duct ligation

AIM: To build up the research models of hepatic fibrosis in mice.METHODS: Inbred wild-type FVB/N mice were either treated with alpha-naphthyl-isothiocyanate (ANIT), allyl alcohol (AA),carbon tetrachloride (CCl4), 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC), and silica, or subjected to common bile duct ligation (CBDL) to induce hepatic injury. Liver biopsies were performed every 4 wk to evaluate hepatic fibrosis over a period of 6 mo. Cumulative cirrhosis and survival curves were constructed by life table method and compared with Wilcoxon test.RESULTS: Under the dosages used, there was neither mortality nor cirrhosis in AA and silica-treated groups. DDC and ANIT caused cirrhosis within 4-12 and 12-24 wk, respectively.Both showed significantly faster cirrhosis induction at high dosages without significant alteration of survival. The duration for cirrhosis induction by CCl4 ranged from 4 to 20 wk, mainly dependent upon the dosage. However, the increase in CCl4dosage significantly worsened survival. Intraperitoneal CCl4administration resulted in better survival in comparison with gavage administration at high dosage, but not at medium and low dosages. After CBDL, all the mice developed liver cirrhosis within 4-8 wk and then died by the end of 16 wk.CONCLUSION: CBDL and administrations of ANIT, CCl4, and DDC ensured liver cirrhosis. CBDL required the least amount of time in cirrhosis induction, but caused shortened lives of mice. It was followed by DDC and ANIT administration with favorable survival. As for CCl4, the speed of cirrhosis induction and the mouse survival depended upon the dosages and the administration route.

胆管阻塞性大鼠肝纤维化模型肝组织MMP2m RNA的表达及复方861抑制作用的研究

观察胆管阻塞性大鼠肝纤维化模型肝组织MMP2mRNA表达以及复方 86 1的抑制作用。给雌性Wis tar大鼠进行胆管内逆行性注入粘合剂加胆管结扎导致胆管阻塞 ,制备胆管阻塞性肝纤维化模型。于术后 7天开始给予不同剂量的复方 86 1灌胃 ,共 4 9天。以RT -PCR法观察肝组织MMP2mRNA表达水平。胆管阻塞性大鼠肝纤维化模型组 (造模 5 6天 )MMP2mRNA为 0 6 33± 0 35 ,明显高于正常对照组 (0 0 2 5± 0 0 18,P <0 0 1) ;复方86 13g/kg、6g/kg、9g/kg组的MMP2mRNA分别为 0 10 7± 0 0 70、0 2 4 8± 0 0 192、0 2 2 2± 0 10 6 ,明显低于模型对照组 (P <0 0 5 )。复方 86 1能够抑制胆管阻塞性大鼠肝纤维化模型肝组织MMP2mRNA表达增高 ,可能是其抗肝纤维化作用的机理之一。

大鼠免疫性与淤胆性肝纤维化发病机制比较

目的大鼠肝脏长期暴露于猪血清后可出现免疫损伤及随后发生的肝纤维化,而胆总管结扎后亦可诱导大鼠发生胆汁淤积性肝损伤,最终发生肝纤维化。这两种损伤后肝脏纤维化的发生、发展及机制均不同。本研究比较这两种不同处理后肝脏损伤的发生发展,探讨炎症与纤维化形成之间的关系。方法(1)通过给大鼠腹腔注射猪血清(IPS)(3ml/kg,1次/周,连续12周)诱导慢性肝纤维化(IPS组)。通过结扎胆总管诱导急性淤胆性肝纤维化(BDL组);(2)采用Knodell组织学评分评价肝脏组织学改变;(3)采用抗αSMA抗体、抗ED1抗体、抗CK7抗体、抗CD45抗体对肝脏进行免疫荧光染色;(4)采用实时定量PCR技术分别检测肝脏中与炎症相关的细胞因子,包括白介素1β(IL-1β)、白介素6(IL-6)、单核细胞趋化蛋白1(MCP-1)、巨噬细胞炎性蛋白1(MIP-1)、肿瘤坏死因子α(TNF-α)、受激活调节正常T细胞表达和分泌因子(RANTES)、转化生长因子β(TGF-β)、血小板衍生生长因子A(PDGF-A);与纤维化相关的细胞因子,包括I型胶原蛋白(col-1)、α平滑肌肌动蛋白(αSMA)、基质金属蛋白酶-9(MMP-9)、金属蛋白酶组织抑制剂-1(TIMP-1)的表达。结果(1)Knodell组织学评分示BDL组汇管区坏死、肝小叶内变性和灶性坏死、汇管区炎症均显著高于IPS组(P<0.01),而肝纤维化评分则低于IPS组(P<0.05);(2)免疫荧光染色显示BDL组肝脏内炎性细胞浸润极为明显,并伴有多量αSMA阳性细胞,IPS组肝脏见较多αSMA阳性细胞,但炎性细胞浸润不明显;(3)实时定量PCR显示与正常对照组肝脏相比,IPS组肝脏中与细胞外基质重建有关的基因表达显著上调;而与炎症相关的细胞因子除RANTES外,均与正常对照组相当(P>0.05)。而BDL组肝脏中所有的与炎症相关的前炎性因子及与纤维化相关的细胞因子均增高(P<0.05)。结论IPS诱导的肝纤维化表现为显著的细胞外基质的重建而不伴明显的炎性细胞浸润,其肝纤维化在无明显活动性炎症的情况下隐袭发展而形成。结扎胆总管诱导的肝纤维化则继发于肝细胞坏死及明显炎症增生。

胆总管结扎术造大鼠肝纤维化模型的研究

目的建立并研究胆总管结扎术造大鼠肝纤维模型的方法。方法 Wista大鼠60只,随机分成假手术组30只和手术组30只,采用胆总管结扎导致胆管阻塞,建立胆管阻塞性肝纤维化模型,分别于术后第5,10,14天将对照组与模型组随机处死10只。分别检测血清学指标,取肝组织做HE染色。结果模型组大鼠体质量均增加,对照组大鼠体质量均降低,肝脾指数明显升高,5天时LN与对照组比较差异有显著性(P<0.01),第10,14天LN、HA与对照组比较差异有显著性(P<0.01),第14天CIV与对照组比较差异有显著性(P<0.01)。模型3组TBIL(μmol/L),DBIL(μmol/L),IBIL(μmol/L)与对照组比较差异差异有显著性(P<0.05)。10天TCHO与对照组比较差异差异有显著性(P<0.05)。AST、ALT均显著升高,模型3组与对照组比较差异有显著性(P<0.05)。HE染色可见同对照组相比,在第5天,模型组的大鼠全部都处于肝纤维化Ⅰ和Ⅱ期阶段;在第10天,模型组的大鼠大部分已经处于肝纤维化Ⅲ期阶段;在第14天,模型组大鼠大部分处于肝纤维化Ⅲ期阶段,一部分处于Ⅳ期阶段。结论胆管阻塞性肝纤维化模型造模周期短,2周左右,自发逆转程度低,肝纤维化形成稳定。

田基黄总黄酮抗胆管结扎所致大鼠肝纤维化的研究

采用胆总管结扎法制备肝纤维化大鼠模型,对田基黄总黄酮抗大鼠肝纤维化作用进行了研究。从造模首日起,灌胃给药,每天给予田基黄总黄酮(9、18、36 mg/kg)1次,4周后,测定血清总胆红素(TBIL)、直接胆红素(DBIL)、谷丙转氨酶(ALT)、谷草转氨酶(AST)、透明质酸(HA)、层粘连蛋白(LN)、Ⅲ型前胶原(PCⅢ)、TNF-α水平及肝组织中超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-Px)活性、丙二醛(MDA)、羟脯氨酸(Hyp)的含量及MMP-1、TIMP-1蛋白的表达。结果表明,田基黄总黄酮能显著降低胆总管结扎诱导的肝纤维化大鼠血清中TBIL、DBIL、ALT、AST、PCⅢ、HA、LN的水平、肝组织中Hyp的含量和TIMP-1蛋白的表达,说明田基黄总黄酮具有良好的抗胆汁性肝纤维化的作用。此外,田基黄总黄酮也能提高肝组织SOD、GSH-Px的活性,降低肝组织MDA及血清中TNF-α的含量。田基黄总黄酮可抑制胆总管结扎诱导的大鼠肝纤维化的形成,其抗肝纤维化作用可能与其抗氧化作用及抗TNF-α的分泌有关。

双氢青蒿素对实验性小鼠肝纤维化的抑制作用

目的探讨双氢青蒿素对胆总管结扎引起的小鼠肝纤维化的抑制作用及其作用机制。方法 50只C57BL-6小鼠随机分为假手术组(n=10)、模型组(n=20)和治疗组(n=20)。模型组:采用胆总管结扎的方法建立小鼠肝纤维化模型;治疗组:在胆总管结扎术后第3周开始以20μg/g的双氢青蒿素灌胃,1次/d,共12次。假手术组:只是分离出胆总管,置线不结扎并缝合腹腔后第3周开始以等剂量的DMSO 0.9%氯化钠溶液灌胃,1次/d,共12次。3组的肝组织切片经HE和天狼猩红染色法观察其组织学病理变化;Western blot方法检测Ⅰ型胶原和α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)蛋白的表达水平。结果经HE和天狼星红染色法结果证实小鼠肝纤维化动物模型的成功建立。模型组小鼠肝内出现灶状坏死,汇管区纤维组织和胆总管明显增生,肝组织中Ⅰ型胶原蛋白和α-SMA含量增加,与假手术组相比其差异有统计学意义(P<0.05)。治疗组经过双氢青蒿素治疗后,小鼠肝内纤维组织增生降低,肝组织中Ⅰ型胶原蛋白和α-SMA含量降低,与模型组比较差异有统计学意义(P<0.05)。结论双氢青蒿素具有抗肝组织纤维化作用,提示可作为一种新型抗肝组织纤维化药物。

胆管阻塞性肝纤维化模型在新生幼犬中的尝试

目的尝试采用新生幼犬制作胆管阻塞性肝纤维化动物模型。方法对新生杂种幼犬进行胆总管双重结扎(胆管结扎组,n=22)或双重结扎并加以切断(胆管切断组,n=22),建立胆管阻塞性肝纤维化模型。观察手术后2、4、6周相关指标的变化情况。结果术后次日所有幼犬均出现尿色深黄,排便陶土样。术后2周时病理组织学观察可见汇管区纤维组织轻度增生;随着梗阻时间延长,局部纤维间隔形成,纤维间隔逐渐向小叶内伸展,局部具有特征性的胆栓形成;最终肝小叶结构破坏,假小叶形成。术后2周,胆管结扎组46%(6/13)的动物出现胆管再通现象,而胆管切断组21%(3/14)的动物出现胆管再通现象,两组比较有显著差异(P<0.001)。结论对幼犬进行单纯胆总管结扎或胆总管结扎后切断都可成功制作出胆管阻塞性肝纤维化模型,但后者成功率更高;幼犬胆管阻塞后形成肝纤维化比较快,自发逆转率低。

血管紧张素Ⅱ1型受体mRNA在胆管阻塞性大鼠肝纤维化模型的表达及复方861的干预作用

研究胆管阻塞性大鼠肝纤维化模型血管紧张素Ⅱ 1型 (AT1)受体mRNA表达以及复方 86 1的治疗作用。给雌性Wistar大鼠进行胆管内逆行性注入硬化剂加胆管结扎导致胆管阻塞 ,制备胆管阻塞性肝纤维化模型。术后随机分为模型组和复方 86 1治疗组 (9g/kg)。并设假手术组作为正常对照组。治疗组于术后 7天开始给予复方 86 1灌胃 ,共 4 9天。以RT -PCR法观察肝纤维化模型肝组织AT1受体mRNA表达水平及复方 86 1的干预作用。胆管阻塞性大鼠肝纤维化模型组AT1受体mRNA为 1 0 38± 0 0 6 4 ,明显高于正常对照组 (0 6 88± 0 0 5 7,P <0 0 1) ;复方86 1组的AT1受体mRNA为 0 70 5± 0 14 2 ,明显低于模型对照组 (P <0 0 1)。胆管阻塞性大鼠肝纤维化模型AT1受体mRNA明显增高 ,提示AT1受体可能参与肝纤维化形成 ,而复方 86 1可使它表达减少。

水飞蓟素、己酮可可碱对大鼠胆管阻塞性肝纤维化的疗效及机制

目的研究水飞蓟素(SIL)、己酮可可碱(PTX)以及两者联合应用的抗纤维化疗效。方法雄性SD大鼠100只,随机分为5组,每组20只,分别为假手术对照组、纤维化模型组、水飞蓟素治疗组(每天50mg/kg)、己酮可可碱治疗组(每天16mg/kg)、联合治疗组。制备胆管堵塞性(BDO)肝纤维化模型。6周后,处死大鼠,取肝组织进行病理学观察并测定肝组织匀浆中的羟脯氨酸(HYP)含量,检测I型前胶原mRNA(procol-ImRNA)和组织金属蛋白酶抑制因子1mRNA(TIMP-1mRNA)的表达水平。检测血清谷丙转氨酶(ALT)、谷草转氨酶(AST)、碱性磷酸酶(ALP)、γ-谷氨酰转肽酶(γ-GT)、总胆红素(TBIL)、直接胆红素(DBIL)、透明质酸(HA)和层连蛋白(LN)水平。结果①与对照组比较,SIL治疗组组织学评分(2.50)比模型组(3.38)明显下降(P<0.01),肝脏HYP减少了35%,Procol-ImRNA下降了35%,TIMP-1mRNA下降了29%。血清HA比模型组降低了30%,LN下降了17%。②PTX治疗组在病理组织学上与水飞蓟素组相似,组织学评分(2.50),HPY比模型组降低38%,Procol-ImRNA减少了33%,TIMP-1mRNA没有减少,并有轻度升高,但无显著性差异。血清HA降低34%,LN下降了13%。③在组织学上,联合治疗组与单独用药相比无明显差别,组织学评分(2.50),与模型组比较,HPY减少了39%,procol-ImRNA降低了33%,但与单独用药组无显著差异。TIMP-1mRNA水平比模型组及PTX组分别降低了25%和27%,并有显著差异,但与SIL组无显著差异。血清HA下降了33%,LN下降了14%,与单独用药组无显著差异。结论单独应用水飞蓟素和己酮可可碱,均改善纤维化程度。两者联合应用与单独用药相比,没有明显差异。这可能TIMP-1mRNA的上调未得到有效对抗有关。

β-catenin在四氯化碳诱导的小鼠肝纤维化过程中的定位、表达及意义

目的探讨肝纤维化发生过程中β-连环蛋白(β-catenin)定位、表达及意义。方法健康雄性昆明小鼠(n=45)随机分为对照组(n=15)与实验组(n=30)。实验组小鼠皮下注射50%四氯化碳(CCl4)-粟米油混合液(6ml/kg),2次∕周,对照组皮下注射同等剂量的粟米油。各组分别于造模1周、4周和8周后取小鼠肝脏,常规制作石蜡切片,Masson染色及Desmin免疫组织化学法观察比较不同组别小鼠肝纤维化病理变化,RT-PCR及免疫组织化学方法检测不同时间点各组小鼠肝组织内β-catenin的表达及定位。结果 Masson染色结果显示,对照组肝汇管区结缔组织内及血管壁有少量细小纤维,实验组小鼠肝脏汇管区和中央静脉及其周围胶原纤维增多;随损伤时间延长纤维增生愈加明显。Desmin免疫组织化学结果显示,各组别均有阳性表达,但损伤组各时间点desmin阳性表达细胞数明显高于对照组(P<0.05或P<0.001)。RT-PCR结果显示,CCl4损伤1周后肝内β-cateninmRNA水平与对照组相比无明显差别(P>0.05),损伤4周及8周β-catenin mRNA水平则明显下降,与对照组相比差异均有显著性(P<0.01)。免疫组织化学结果显示,对照组β-catenin弱表达于肝细胞膜及胆管上皮细胞膜和胞质,而损伤组β-catenin阳性反应主要定位于肝内增生的细胞团及新生胆管上皮细胞质,各组间积分吸光度值差别有显著性(P<0.01)。结论 CCl4诱导的肝纤维化过程中β-catenin mRNA表达与蛋白表达不同步,阳性表达细胞主要为新生的细胞团及胆管上皮细胞。

消黄方对大鼠胆汁淤积性肝纤维化的干预作用

目的:观测消黄方对胆管结扎大鼠肝功能及组织病理学的干预作用。方法:Wistar大鼠30只,随机分为假手术组和胆管结扎组(BDL)。BDL大鼠在2周末随机分为消黄方组和模型组,第3周开始消黄方组每天灌胃给药,假手术组和BDL组灌胃等量生理盐水,共3周。5周末处死全部大鼠,获取血清和肝组织样本。结果:与假手术组相比,模型组血清谷草转氨酶(ALT),谷丙转氨酶(AST)和碱性磷酸酶(ALP)活性及总胆红素(TBIL)含量显著上升(P<0.05或0.01),白蛋白(Alb)含量显著下降(P<0.01);与模型组相比,消黄方显著降低ALP活性和TBIL含量,显著抑制Alb含量的降低(P<0.05或0.01),降低肝组织Hyp含量(P<0.05),同时病理学显示消黄方组肝纤维化程度也显著减轻。结论:消黄方能显著抑制胆汁淤积性肝纤维化的形成。

大鼠胆管阻塞性肝纤维化模型的建立

目的 建立大鼠胆管阻塞性肝纤维化模型 ,为肝纤维化机制和治疗研究提供基础。方法 雌性Wistar大鼠进行胆管内逆行性注入N 丁基 2 氰基丙烯酸盐加胆管结扎导致胆管阻塞 ,建立胆管阻塞性肝纤维化模型。观察手术后第 2 8d和 5 6d不同时间相关指标的改变。结果 胆管阻塞性肝纤维化模型ALT、AST、HA、LN均明显升高 ;病理组织学可见弥漫增生的胆小管 ,大量的胶原沉积在胆小管周围 ,肝细胞与增生的胆小管相比明显减少。结论 胆管阻塞性肝纤维化模型造模周期短 ,自发逆转率低 ,肝纤维化形成稳定 。

水飞蓟素对实验性肝纤维化的疗效及其作用机制的研究

目的:观察水飞蓟素(Silymarin,SIL)在胆管阻塞性肝纤维化大鼠模型(bile duct occlusion, BDO)中的抗纤维化疗效。方法:雄性SD大鼠60只,随机分为3组,每组20只,分别为假手术对照组、纤维化模型组、水飞蓟素治疗组(50 mg·kg-1·d-1)。制备胆管阻塞性肝纤维化模型。用药6周后,处死大鼠,取肝组织进行病理学观察。测定肝组织羟脯氨酸(HYP)含量。检测I型前胶原(procol-I)mRNA和组织金属蛋白酶抑制因子I(TIMP-1)mRNA表达水平及血清透明质酸、层连蛋白、转氨酶、碱性磷酸酶、γ-谷氨酰转肽酶、胆红素水平。实验结果统计采用SPSS软件进行分析,P<0.05为差异有显著性意义。结果:与对照组比较,水飞蓟素治疗组的组织学评分(2.50)比模型组(3.38)明显下降(P<0.01)。肝脏HYP减少35%。procol-I mRNA和TIMP-1 mRNA分别下降35%和29%。血清HA降低30%,LN下降17%。以上差异均有统计学意义。结论:水飞蓟素抑制procol-I mRNA、TIMP-1mRNA表达,减少肝组织胶原含量,改善肝纤维化程度。