Sallés F J, Schechter N, Strickland S
Department of Molecular and Cellular Pharmacology, State University of New York, Stony Brook 11794.
EMBO J. 1990 Aug;9(8):2471-7. doi: 10.1002/j.1460-2075.1990.tb07425.x.
The use of purified piscine plasminogen in a chromogenic solution assay enabled us to detect plasminogen activator (PA) activity in crude homogenates of goldfish optic nerve following nerve injury. In contrast, no activity was detected in the homogenates of uninjured nerve. Under conditions allowing regeneration of the optic axons (optic nerve crush), PA activity peaked 8 days after crush, and decreased to undetectable levels by 60 days. Under conditions allowing only degeneration of the axons (enucleation), the activity peaked at 8 days but decreased more rapidly. Casein zymography of samples after fractionation in SDS-PAGE showed that PA activity migrated as a doublet at Mr = 60-65 kd. Using this assay, activity was also observed in uninjured control nerves. This plasminogen-dependent activity migrated as three bands of higher molecular weight (Mr = 75, 95 and 120 kd) and was undetectable in solution assays of unfractionated extracts, suggesting complex formation with an inhibitor(s). Fibrin overlay assay of retinal explants and isolated primary cells in culture suggest that the goldfish PA is associated with the glial cells of the goldfish visual pathway.
在发色底物溶液测定中使用纯化的鱼纤溶酶原,使我们能够检测金鱼视神经损伤后粗匀浆中的纤溶酶原激活物(PA)活性。相比之下,在未损伤神经的匀浆中未检测到活性。在允许视神经轴突再生的条件下(视神经挤压),PA活性在挤压后8天达到峰值,并在60天时降至无法检测的水平。在仅允许轴突退变的条件下(眼球摘除),活性在8天达到峰值,但下降得更快。SDS-PAGE分级分离后的样品进行酪蛋白酶谱分析表明,PA活性以Mr = 60-65 kd的双条带形式迁移。使用该测定法,在未损伤的对照神经中也观察到活性。这种纤溶酶原依赖性活性以三条较高分子量的条带(Mr = 75、95和120 kd)形式迁移,并且在未分级提取物的溶液测定中无法检测到,这表明与一种或多种抑制剂形成了复合物。视网膜外植体和培养的分离原代细胞的纤维蛋白覆盖测定表明,金鱼PA与金鱼视觉通路的神经胶质细胞相关。