Nagasaki A, Gotoh T, Takeya M, Yu Y, Takiguchi M, Matsuzaki H, Takatsuki K, Mori M
Department of Molecular Genetics, Kumamoto University School of Medicine, Japan.
J Biol Chem. 1996 Feb 2;271(5):2658-62. doi: 10.1074/jbc.271.5.2658.
Nitric oxide (NO) is synthesized from arginine by nitric oxide synthase (NOS), and citrulline which is generated can be recycled to arginine by argininosuccinate synthetase (AS) and argininosuccinate lyase (AL). Rats were injected with bacterial lipopolysaccharide (LPS), and expression of the inducible isoform of NOS (iNOS), AS, and AL was analyzed. In RNA blot analysis, iNOS mRNA was undetectable before the LPS treatment but was induced by LPS in the lung, heart, liver, and spleen, and less strongly in the skeletal muscle and testis. AS mRNA was induced in the lung and spleen, and AL mRNA was weakly induced in these tissues. AS and AL mRNAs were abundant in the control liver and remained unchanged after the treatment. Kinetic studies showed that iNOS mRNA increased rapidly in both spleen and lung, reached a maximum 2-5 h after the treatment, and decreased thereafter. On the other hand, AS mRNA increased more slowly and reached a maximum in 6-12 h (by about 10-fold in the spleen and 2-fold in the lung). AL mRNA in the spleen and lung increased slowly and remained high up to 24 h. In immunoblot analysis, increase of iNOS protein was evident in the lung, liver, and spleen, and there was an increase of AS protein in the lung and spleen. In immunohistochemical analysis, macrophages in the spleen that were negative for iNOS and AS before LPS treatment were strongly positive for both iNOS and AS after this treatment. As iNOS, AS, and AL were coinduced in rat tissues and cells, citrulline-arginine recycling seems to be important in NO synthesis under the conditions of stimulation.
一氧化氮（NO）由一氧化氮合酶（NOS）催化精氨酸合成，生成的瓜氨酸可通过精氨琥珀酸合成酶（AS）和精氨琥珀酸裂解酶（AL）再循环为精氨酸。给大鼠注射细菌脂多糖（LPS），分析诱导型NOS（iNOS）、AS和AL的表达。在RNA印迹分析中，LPS处理前未检测到iNOS mRNA，但在肺、心脏、肝脏和脾脏中被LPS诱导，在骨骼肌和睾丸中诱导较弱。AS mRNA在肺和脾脏中被诱导，AL mRNA在这些组织中被弱诱导。AS和AL mRNA在对照肝脏中丰富，处理后保持不变。动力学研究表明，iNOS mRNA在脾脏和肺中均迅速增加，处理后2 - 5小时达到最大值，此后下降。另一方面，AS mRNA增加较慢，在6 - 12小时达到最大值（脾脏中约增加10倍，肺中增加2倍）。脾脏和肺中的AL mRNA增加缓慢，直至24小时仍保持高水平。在免疫印迹分析中，肺、肝脏和脾脏中iNOS蛋白明显增加，肺和脾脏中AS蛋白增加。在免疫组织化学分析中，LPS处理前iNOS和AS阴性的脾脏巨噬细胞在处理后iNOS和AS均呈强阳性。由于iNOS、AS和AL在大鼠组织和细胞中共同诱导，在刺激条件下瓜氨酸 - 精氨酸循环似乎在NO合成中起重要作用。
