Hanumegowda Umesh M, Copple Bryan L, Shibuya Masabumi, Malle Ernst, Ganey Patricia E, Roth Robert A
Department of Pharmacology and Toxicology, Michigan State University, East Lansing, Michigan 48824, USA.
Toxicol Sci. 2003 Nov;76(1):237-46. doi: 10.1093/toxsci/kfg222. Epub 2003 Sep 11.
Monocrotaline (MCT) is a pyrrolizidine alkaloid that causes liver injury in animals. In rats, injury is characterized by sinusoidal endothelial cell (SEC) damage and centrilobular parenchymal cell necrosis. Loss of endothelium is a possible outcome of the action of matrix metalloproteinases (MMPs), specifically MMP-9 from neutrophils and SECs and MMP-2 from SECs, on basement membrane collagen. Accordingly, the dynamics of MMPs in MCT-induced SEC damage were studied. Rats were treated with MCT (300 mg/kg, ip), and livers were collected at 8, 12, and 18 h. Immunofluorescence analysis of frozen sections of livers from MCT-treated rats revealed a progressive reduction in basement membrane heparan sulfate proteoglycan and collagen IV. A time-dependent increase in total type IV collagenase activity and MMP-9 content occurred in the livers of MCT-treated rats, as measured by fluorescent collagenase activity assay and gelatin zymography, respectively. Progressive neutrophil accumulation and activation in the liver after MCT treatment were demonstrated by an increased activity of myeloperoxidase and pronounced staining for hypochlorite-modified proteins generated via the myeloperoxidase-hydrogen peroxide-halide system. However, neutrophil depletion did not protect against MCT-induced SEC injury. Treatment of NP-26 cells, a sinusoidal endothelial cell line, with MCT resulted in dose-dependent release of MMP-9 from the cells. The results demonstrate the degradation of basement membrane components with a concurrent increase in the amount and activity of MMP-9, likely originating from sinusoidal endothelial cells, neutrophils, and probably other cell types. This suggests the possibility of a role for MMPs in the SEC detachment and loss that occurs during MCT hepatotoxicity.
野百合碱（MCT）是一种吡咯里西啶生物碱，可导致动物肝脏损伤。在大鼠中，损伤的特征是肝血窦内皮细胞（SEC）损伤和小叶中央实质细胞坏死。内皮细胞的丧失可能是基质金属蛋白酶（MMPs）作用的结果，具体而言，中性粒细胞和SEC产生的MMP-9以及SEC产生的MMP-2作用于基底膜胶原蛋白。因此，研究了MMPs在MCT诱导的SEC损伤中的动态变化。给大鼠腹腔注射MCT（300 mg/kg），并在8、12和18小时收集肝脏。对MCT处理的大鼠肝脏冰冻切片进行免疫荧光分析，结果显示基底膜硫酸乙酰肝素蛋白聚糖和IV型胶原蛋白逐渐减少。通过荧光胶原酶活性测定和明胶酶谱法分别测定，MCT处理的大鼠肝脏中IV型胶原酶总活性和MMP-9含量呈时间依赖性增加。MCT处理后，肝脏中中性粒细胞逐渐积聚并活化，髓过氧化物酶活性增加以及通过髓过氧化物酶-过氧化氢-卤化物系统产生的次氯酸盐修饰蛋白染色明显可证明这一点。然而，中性粒细胞耗竭并不能预防MCT诱导的SEC损伤。用MCT处理肝血窦内皮细胞系NP-26细胞，导致细胞中MMP-9呈剂量依赖性释放。结果表明基底膜成分降解，同时MMP-9的量和活性增加，MMP-9可能源自肝血窦内皮细胞、中性粒细胞以及可能的其他细胞类型。这表明MMPs在MCT肝毒性期间发生的SEC脱离和丧失中可能发挥作用。
