环氧合酶-2在四氯化碳诱导肝硬化大鼠肝窦毛细血管化形成中的作用

Effects of cyclooxygenase-2 on sinusoidal capillarization in cirrhotic rats induced by carbon tetrachloride

目的观察环氧合酶-2（COX-2）在实验性肝硬化大鼠肝窦毛细血管化形成中的作用。方法腹腔注射CCl4每周2次共8周诱导雄性SD大鼠肝硬化模型。将SD大鼠分成3组：正常对照组（n=10）、模型对照组（n=15）和罗非昔布治疗组（10mg·kg^-1·d^-1，n=15）。光镜下观察肝组织标本，电镜观察肝窦超微结构改变。用Western印迹和免疫组化法检测基底膜蛋白主要成分层粘连蛋白（LN）和Ⅳ型胶原，同时通过Ⅷ因子相关抗原（vWF）免疫组化标记微血管生成密度。结果与模型对照组相比，罗非昔布干预治疗能减少肝纤维化面积（分别为30．7±8．9和23．5±6．5，P〈0．05）。光镜及电镜提示，在模型对照组可见肝窦内皮细胞窗孔减少、缩小，有完整的基底膜形成，Disse腔隙内有大量的胶原纤维沉积，罗非昔布组上述病变有所减轻。随着肝硬化的形成，肝组织微血管密度明显升高，罗非昔布组肝组织微血管密度（6．4±0．7）较模型对照组（11．3±1．6）明显降低（P〈0．01）。肝硬化时肝组织表达Ⅳ型胶原和LN蛋白明显增加（分别为3．8±0．4和3．7±0．5），罗非昔布能降低Ⅳ型胶原和LN的表达（分别为3．0±0．5和3．0±0．5；与模型对照组相比两者均为P〈0．05）。结论在CCl4诱导的肝硬化大鼠模型中，早期给予罗非昔布能有效改善肝窦毛细血管化。

Objective To investigate the role of cyclooxygenase-2 （COX-2） in sinusoidal capillarization in liver cirrhotic rats. Methods The SD rats were intraperitoneally injected with carbon tetrachloride （CCl4） twice a week for 8 weeks to induce liver cirrhosis. The rats were randomly divided into three groups; normal control group （n= 10）, model control group （n=15） and rofecoxib treated group （received 10 mg/kg of rofecoxib daily, n= 15）. Liver histopathology was examined by light microscopy, and sinusoidal ultrastructure was observed by transmission electron microscopy. Furthermore, the level of basement membrane proteins （collagen type Ⅳ, laminin） and their localizations in liver were determined by Western blotting and immunohistochemistry, respectively, and the microvessel density was detected following vWF （yon Willebrand factor） immunolabeling on liver tissue sections. Results Fibrotic areas were reduced in rofecoxib treated group compared with that in model group （30.7±8.9 vs 23.5±6.5,P〈0.05）. The light and electron microscopy showed that the pathologic changes including loss or reduction in number of sinusoidal endothelial fenestrate, the accumulation of extracellular matrix in the Disse＇s space and development of subendothelial basal lamina （basement membrane formation） were more severe in model group than those in rofecoxib treated group. Compared with model group, administration of rofecoxib resulted in significant decrease in microvessel density （11.3 ± 1.6 vs. 6.4 ±0. 7, P〈0. 01）. Rofecoxib could significantly decrease the expression of type Ⅳ collagen and laminin at protein levels （3.0±0.5 and 3.0±0.5, respectively） when compared with model group （3.8 ± 0.4 and 3.7 ±0.5, respectively）. Conelusion The results indicate that early administration of rofecoxib may reduce sinusoidal capillarization.