Siller-López Fernando, Sandoval Ana, Salgado Silvia, Salazar Adriana, Bueno Miriam, Garcia Jesus, Vera Jose, Gálvez Javier, Hernández Iván, Ramos Martha, Aguilar-Cordova Estuardo, Armendariz-Borunda Juan
Institute for Molecular Biology in Medicine and Gene Therapy, Centro Universitario de Ciencias de la Salud, University of Guadalajara, Mexico.
Gastroenterology. 2004 Apr;126(4):1122-33; discussion 949. doi: 10.1053/j.gastro.2003.12.045.
BACKGROUND & AIMS: An extrahepatic human neutrophil collagenase complementary DNA (matrix metalloprotease-8) cloned in an adenovirus vector was used as a therapeutic agent in cirrhosis.
A high titer of clinical-grade AdMMP8 was obtained.
HeLa cells transduced with AdMMP8 expressed recombinant matrix metalloprotease-8 messenger RNA and matrix metalloprotease-8 protein. Matrix metalloprotease-8 in culture sups showed enzymatic activity against native collagen type I, which was inhibited by ethylenediaminetetraacetic acid, 1,10-phenanthroline, and tissue inhibitor of metalloprotease-1. In vivo transduction showed matrix metalloprotease-8 activity, and studies to establish the efficacy of this characterized vector were performed in CCl(4) and bile duct-ligated cirrhotic rats. Transduction with 3 x 10(11) viral particles per kilogram resulted in hepatic detection of both messenger RNA and protein matrix metalloprotease-8. A consistent response in fibrosis reversal was observed in CCl(4) rats. Liver fibrosis in bile duct-ligated cirrhotic animals was decreased in 45%, along with diminished hydroxyproline content, after AdMMP8 treatment. The expression of matrix metalloprotease-2 and matrix metalloprotease-3 was up-regulated in AdMMP8 rats. Free tissue inhibitor of metalloprotease-1, as an indirect measurement of active uncomplexed matrix metalloproteases, was also increased in the AdMMP8 groups. Transforming growth factor-beta messenger RNA was diminished, and matrix metalloprotease-9 and hepatocyte growth factor increased. Treatment in both models correlated with improvements in ascites, functional hepatic tests, and gastric varices, indicating diminished intrahepatic blood pressure in animals injected with AdMMP8.
Therefore, therapy with the matrix metalloprotease-8 gene is promising for use in a clinical setting.
克隆于腺病毒载体的人肝外中性粒细胞胶原酶互补DNA（基质金属蛋白酶-8）被用作肝硬化的治疗药物。
获得了高滴度的临床级AdMMP8。
用AdMMP8转导的HeLa细胞表达重组基质金属蛋白酶-8信使核糖核酸和基质金属蛋白酶-8蛋白。培养上清液中的基质金属蛋白酶-8对天然I型胶原显示出酶活性，该活性被乙二胺四乙酸、1,10-菲咯啉和金属蛋白酶-1组织抑制剂所抑制。体内转导显示出基质金属蛋白酶-8活性，并在四氯化碳和胆管结扎的肝硬化大鼠中进行了研究以确定这种特性载体的疗效。每千克用3×10¹¹个病毒颗粒进行转导，导致肝脏中检测到信使核糖核酸和基质金属蛋白酶-8蛋白。在四氯化碳大鼠中观察到纤维化逆转的一致反应。AdMMP8治疗后，胆管结扎的肝硬化动物的肝纤维化减少了45%，同时羟脯氨酸含量降低。AdMMP8大鼠中基质金属蛋白酶-2和基质金属蛋白酶-3的表达上调。作为活性未复合基质金属蛋白酶的间接测量指标，游离金属蛋白酶-1组织抑制剂在AdMMP8组中也增加。转化生长因子-β信使核糖核酸减少，基质金属蛋白酶-9和肝细胞生长因子增加。两种模型中的治疗都与腹水、肝功能试验和胃静脉曲张的改善相关，表明注射AdMMP8的动物肝内血压降低。
因此，基质金属蛋白酶-8基因治疗在临床应用中具有前景。
