Zhu W, Diwan A D, Lin J H, Murrell G A
Orthopedic Research Institute, St George Hospital, University of New South Wales, Sydney, Australia.
J Bone Miner Res. 2001 Mar;16(3):535-40. doi: 10.1359/jbmr.2001.16.3.535.
We have shown previously that nitric oxide (NO) has regulatory effects on fracture healing. Our aim here was to investigate the temporal expression patterns of the three NO synthase (NOS) isoforms that are responsible for the generation of NO by semiquantitative competitive polymerase chain reaction (PCR) and immunoblot analysis after femoral fractures in rats. We found that 4 days after fracture, there were increases in the levels of messenger RNA (mRNA) for all three NOS isoforms, with peaks for the inducible NOS (iNOS; 35-fold increase, p < 0.05) at day 4, the endothelial NOS (eNOS; 5-fold increase, p < 0.05) at day 7, and the neuronal NOS (bNOS; 16-fold increase, p < 0.05) at day 21. At a protein level, the time course expression of NOS isoforms was consistent with the results of those at the mRNA level. In addition, we have previously reported a 2.5-fold increase in NOS activity detected by [3H]arginine to [3H]citrulline conversion at day 15 compared with that at day 4 after fracture. The findings that the expression of NOS isoforms during fracture healing is type specific and time dependent are important and may have clinical applications in the regulation of bone repair by NOS inhibitors or stimulators at different stages after injury.
我们之前已经表明，一氧化氮（NO）对骨折愈合具有调节作用。我们在此的目的是通过半定量竞争性聚合酶链反应（PCR）和免疫印迹分析，研究大鼠股骨骨折后负责生成NO的三种一氧化氮合酶（NOS）亚型的时间表达模式。我们发现骨折后4天，所有三种NOS亚型的信使核糖核酸（mRNA）水平均升高，其中诱导型NOS（iNOS；增加35倍，p<0.05）在第4天达到峰值，内皮型NOS（eNOS；增加5倍，p<0.05）在第7天达到峰值，神经元型NOS（nNOS；增加16倍，p<0.05）在第21天达到峰值。在蛋白质水平上，NOS亚型的时间进程表达与mRNA水平的结果一致。此外，我们之前报道过，与骨折后第4天相比，在第15天通过[3H]精氨酸向[3H]瓜氨酸转化检测到的NOS活性增加了2.5倍。骨折愈合过程中NOS亚型的表达具有类型特异性和时间依赖性，这一发现很重要，并且可能在损伤后不同阶段通过NOS抑制剂或刺激剂调节骨修复方面具有临床应用价值。
