Pizzi Michael A, Elam John S
Program in Neuroscience, Department of Biological Science, Florida State University, Tallahassee, Florida 32306-3050, USA.
Neurochem Res. 2004 Apr;29(4):719-28. doi: 10.1023/b:nere.0000018842.90981.41.
Regeneration of goldfish optic nerve axons is accompanied by a major increase in axonally transported proteoglycans (21). To identify specific proteoglycans increased during regeneration, we have used proteoglycan preparations from regenerating goldfish optic tracts to produce monoclonal antibodies. Western blot analysis shows a 28-kD antigen reacting with our 1G4/G5 antibody is present in optic tract 21 days after nerve crush, but absent in nonregenerating tract. Treatment with chondroitinase AC removes IG4/G5 immunostaining of the 28-kD molecule. An anti-CS antibody further confirmed this molecule as a chondroitin sulfate proteoglycan. A slightly smaller core protein following chondroitinase AC treatment indicates a low level of glycosylation. The N-terminal amino acid sequence of its core protein is not similar to any known proteoglycans. The CSPG sediments through 1.4 M sucrose, indicative of an extracellular matrix localization. It is expressed during the outgrowth of regenerating axons. Cut nerve and retinal explant studies demonstrate that the 1G4/G5 CSPG is not axonal, suggesting a glial localization.
金鱼视神经轴突的再生伴随着轴突运输的蛋白聚糖的显著增加(21)。为了鉴定再生过程中增加的特定蛋白聚糖,我们使用了来自再生金鱼视束的蛋白聚糖制剂来制备单克隆抗体。蛋白质印迹分析表明,与我们的1G4/G5抗体发生反应的一种28-kD抗原在神经挤压后21天存在于视束中,但在非再生视束中不存在。用软骨素酶AC处理可消除28-kD分子的IG4/G5免疫染色。一种抗CS抗体进一步证实该分子是一种硫酸软骨素蛋白聚糖。软骨素酶AC处理后核心蛋白略小,表明糖基化水平较低。其核心蛋白的N端氨基酸序列与任何已知的蛋白聚糖都不相似。该硫酸软骨素蛋白聚糖在1.4 M蔗糖中沉降,表明其定位于细胞外基质。它在再生轴突生长过程中表达。切断神经和视网膜外植体研究表明,1G4/G5硫酸软骨素蛋白聚糖不是轴突性的,提示其定位于神经胶质细胞。