| 摘要: |
| 目的:采用α- 萘异硫氰酸酯(ANIT) 多次灌胃给药制备慢性胆汁淤积肝纤维化大鼠模型，观察绿原酸对该模型胆管及胶原增生的影响，探讨其防治机制。方法:健康成年SPF 级Wistar 雄性大鼠40 只，随机分为5 组:对照组、模型组、熊去氧胆酸(UDCA)组、绿原酸低剂量组和绿原酸高剂量组，每组8 只。除对照组外，其余4 组均以ANIT(80 mg/kg)灌胃5 周、每周3 次，制备慢性胆汁淤积模型，同时UDCA 组、绿原酸低剂量组和绿原酸高剂量组，分别给予UDCA(90mg/Kg)、绿原酸(50 mg/Kg 和100 mg/Kg)灌胃，每日1 次，连续5 周。治疗5 周结束后，连同对照组、模型组，共5 组动物，麻醉状态下，经腹主动脉取血，全自动生化分析仪检测血清中总胆红素(TBIL)、结合胆红素(DBIL)、间接胆红素(IBIL)以及碱性磷酸酶(ALP)水平。切取部分肝组织，中性甲醛固定，进行HE 染色、天狼星红染色，观察评价肝组织胆汁淤积、胆管增生及胶原增生等病理改变情况；另取部分肝组织，匀浆提取总蛋白，Western blot 法检测肝星状细胞活化标志分子-α-平滑肌肌动蛋白(α-SMA)表达水平。结果:与正常组比，模型组大鼠血清TBIL、DBIL、IBIL、ALP 水平显著升高(均P <0.01)；与模型组比，绿原酸高、低剂量组以及UDCA 组血清TBIL、DBIL、IBIL 水平显著降低(P<0.05)，ALP 有下降趋势，但差异无显著性(P>0.05)。模型组HE 染色显示肝组织慢性炎性细胞浸润明显，汇管区胆管增生、扩张显著；绿原酸高、低剂量组及UDCA 组肝慢性炎性反应及汇管区胆管增生反应显著减轻。天狼星红染色显示正常组大鼠肝组织胶原表达稀少，而模型组汇管区胶原表达显著增强，偏振光下以亮红黄色的I 型胶原为主；与模型组比，绿原酸高、低剂量组以及UDCA 组肝组织胶原染色均显著减轻，偏振光下以呈绿色的III 型胶原为主。Western blot 法检测显示，模型组大鼠肝组织中α-SMA 表达水平显著升高(P <0.01)，与模型组比，绿原酸高、低剂量组以及UDCA 组肝组织α-SMA 表达显著降低(均P <0.05)。结论:绿原酸可显著改善慢性胆汁淤积大鼠模型的肝功能损伤、有效抑制肝星状细胞活化、减少以I 型胶原为主的胶原产生、减轻慢性胆汁淤积所致的汇管区胆管增生，从而发挥抗肝纤维化作用。 |
| 关键词: 绿原酸 α- 萘异硫氰酸酯 慢性胆汁淤积 α- 平滑肌肌动蛋白 胶原 |
| DOI：10.3969/j.issn.1007-6948.2020.03.005 |
| 投稿时间：2020-01-11 |
| 基金项目: |
|
| Effect of Chlorogenic Acid on the Proliferation of Bile Duct and Collagen in Rats with Liver Fibrosis Induced by Chronic Cholestasis |
| TANG Li-ming,Li Wan-hua,SONG Ning |
|
| Abstract: |
| Objective To observe the effect of chlorogenic acid on the proliferation of bile duct and collagen in rats with chronic cholestasis and to explore the mechanism of prevention and treatment. Methods Forty adult male Wistar rats were randomly divided into five groups: control group, model group, UDCA group,low dose group and high dose group. In addition to the control group, the other four groups were treated with ANIT (80 mg/kg) for 5 weeks and 3 times a week to prepare the chronic cholestasis model. At the same time,UDCA group, low dose group and high dose group were given UDCA (90 mg/kg), chlorogenic acid (50 mg/kgand 100 mg/kg) for 5 weeks. At the end of 5 weeks, 5 groups of animals, including control group and model group, were taken blood from abdominal aorta under anesthesia, and the levels of TBIL, DBIL, ibil and ALP in serum were measured by automatic biochemical analyzer. Partial liver tissues were cut, fixed with neutral formaldehyde, stained with he and Sirius red,and the pathological changes of cholestasis, bile duct hyperplasia and collagen hyperplasia were observed and evaluated. The total protein was extracted from other liver tissues, and the expression level of α-smooth muscle actin (α-SMA) was detected by Western blot. Results compared with the normal group, the levels of TBIL, DBIL, IBIL and ALP in the model group were signi?cantly higher (P<0.01); compared with the model group, the levels of TBIL, DBIL and IBIL in the high and low dose chlorogenic acid group and UDCA group were signi?cantly lower (P<0.05), and ALP had a downward trend, but there was no signi?cant difference (P>0.05). HE staining of the model group showed that the chronic inflammatory cell infiltration in the liver tissue was obvious, the proliferation and expansion of bile duct in the portal area were signi?cant; the chronic in?ammatory response and the proliferation of bile duct in the portal area were signi?cantly reduced in the high and low dose groups of chlorogenic acid and UDCA groups. Sirius red staining showed that the expression of collagen in the normal group was rare, while that in the model group was signi?cantly enhanced. Under polarized light, collagen I was mainly bright red and yellow. Compared with the model group, collagen staining in the high and low dose groups of chlorogenic acid and UDCA group was signi?cantly reduced, while collagen III was mainly green under polarized light. Western blot showed that the expression level of α-SMA in the liver tissue of the model group increased significantly (P<0.01), compared with the model group, the expression of α - SMA in the liver tissue of the high dose group, the low dose group and the UDCA group decreased signi?cantly (P<0.05). Conclusion Chlorogenic acid can signi?cantly improve the liver function damage, effectively inhibit the activation of hepatic stellate cells, reduce the production of collagen based on type I collagen, and reduce the proliferation of bile duct in the portal area caused by chronic cholestasis, so as to play an anti ?brosis role. |
| Key words: Chlorogenic acid α-naphthalene isothiocyanate chronic cholestasis α-smooth muscle actin collagen |
