Van der Zee C E, Nielander H B, Vos J P, Lopes da Silva S, Verhaagen J, Oestreicher A B, Schrama L H, Schotman P, Gispen W H
Division of Molecular Neurobiology, Rudolf Magnus Institute, University of Utrecht, The Netherlands.
J Neurosci. 1989 Oct;9(10):3505-12. doi: 10.1523/JNEUROSCI.09-10-03505.1989.
Recently it has been shown that B-50 is identical to the neuron-specific, growth-associated protein GAP43. The present study reports on the fate of B-50/GAP43 mRNA and B-50/GAP43 protein, determined by radioimmunoassay, in a rat model of peripheral nerve regeneration (sciatic nerve crush) over a period of 37 and 312 d, respectively. Moreover, the effects of repeated subcutaneous injection of the neurotrophic peptide Org.2766 (an ACTH4-9 analog) and of a conditioning lesion on B-50/GAP43 protein levels in the regenerating nerve and dorsal root ganglia (DRG) were investigated. Both treatments enhanced the functional recovery as evidenced by a foot-flick withdrawal test. Immunocytochemical analysis using antineurofilament antibodies revealed a peptide-induced increase in the number of outgrowing sprouts in the sciatic nerve. Both the peptide and the conditioning lesion amplified the crush lesion-induced increase in B-50 protein content in the nerve as determined by radioimmunoassay. B-50 protein levels seem to correlate proportionally with the number of sprouts. In the DRG of the crushed sciatic nerve, the time course of B-50 expression was studied. B-50 mRNA was quantified from Northern blots. A linear increase up to 10 times the basal level of B-50 mRNA was observed 2 d postsurgery, followed by a gradual decline to normal levels at day 37. The first significant rise in B-50 mRNA level became apparent between 8 and 16 hr after placement of the crush lesion. The first significant rise in B-50 protein level occurred 40 hr after the crush lesion, reaching a plateau of 3 times the basal level between day 6 and 20. B-50 protein levels in DRG cell bodies remained elevated up to 60 d after crush, a period much longer than that observed for B-50 mRNA. Thus, during a later phase of peripheral axonal regeneration, the presence of B-50 appears to be prolonged, probably by an increase in half-life and not so much by enhanced transcription. Treatment with Org.2766 did not affect the B-50/GAP43 levels in DRG cell bodies during the first 6 d following crush. Conditioning lesion resulted in a DRG B-50/GAP43 protein amount at the same level as in rats 14 d after the test lesion. B-50/GAP43 levels in DRG are probably influenced by the rapid axonal transport of the protein, as has been reported by others.
最近研究表明，B - 50与神经元特异性生长相关蛋白GAP43相同。本研究报告了在大鼠外周神经再生模型（坐骨神经挤压伤）中，分别于37天和312天期间，通过放射免疫分析测定的B - 50/GAP43 mRNA和B - 50/GAP43蛋白的变化情况。此外，还研究了反复皮下注射神经营养肽Org.2766（一种促肾上腺皮质激素4 - 9类似物）以及预处理损伤对再生神经和背根神经节（DRG）中B - 50/GAP43蛋白水平的影响。两种处理均增强了功能恢复，这通过甩足反射试验得以证明。使用抗神经丝抗体的免疫细胞化学分析显示，该肽可诱导坐骨神经中长出的新芽数量增加。放射免疫分析测定结果表明，该肽和预处理损伤均放大了挤压损伤诱导的神经中B - 50蛋白含量的增加。B - 50蛋白水平似乎与新芽数量成比例相关。在挤压伤坐骨神经的DRG中，研究了B - 50的表达时间进程。从Northern印迹定量分析B - 50 mRNA。术后2天观察到B - 50 mRNA水平线性增加至基础水平的10倍，随后逐渐下降至第37天的正常水平。挤压损伤后8至16小时之间，B - 50 mRNA水平首次出现显著升高。挤压损伤后40小时，B - 50蛋白水平首次出现显著升高，在第6天至20天之间达到基础水平3倍的平台期。挤压伤后长达60天，DRG细胞体中的B - 50蛋白水平一直保持升高，这一时期比观察到的B - 50 mRNA的时期长得多。因此，在外周轴突再生的后期阶段，B - 50的存在似乎延长，可能是通过半衰期的延长，而不是转录增强。挤压伤后的前6天，用Org.2766处理并未影响DRG细胞体中的B - 50/GAP43水平。预处理损伤导致DRG中的B - 50/GAP43蛋白量与试验损伤后14天的大鼠处于同一水平。正如其他人所报道的，DRG中的B - 50/GAP43水平可能受该蛋白快速轴突运输的影响。
