Hammermann R, Hirschmann J, Hey C, Mössner J, Folkerts G, Nijkamp F P, Wessler I, Racké K
Institute of Pharmacology and Toxicology, University of Bonn, Bonn; Department of Pharmacology, University of Mainz, Mainz, Germany.
Am J Respir Cell Mol Biol. 1999 Aug;21(2):155-62. doi: 10.1165/ajrcmb.21.2.3574.
Eosinophil-derived cationic proteins play an essential role in the pathogenesis of bronchial asthma. We tested whether cationic proteins interfere with the cationic amino-acid transport in alveolar macrophages (AMPhi) and tracheal epithelial cells, and whether L-arginine-dependent pathways were affected. The effect of cationic polypeptides on cellular uptake of [(3)H]-L-arginine, nitrite accumulation, and the turnover of [(3)H]-L-arginine by nitric oxide (NO) synthase and arginase (formation of [(3)H]-L-citrulline and [(3)H]-L-ornithine, respectively) were studied. Poly-L-arginine reduced [(3)H]-L-arginine uptake in rat AMPhi and tracheal epithelial cells in a concentration-dependent manner (at 300 microgram/ml by 70%). Poly-L-lysine, protamine, and major basic protein (each up to 300 microgram/ml) tested in rat AMPhi inhibited [(3)H]-L-arginine uptake by 35 to 50%. During 6 h incubation in amino acid-free Krebs solution, rat AMPhi, precultured in the absence or presence of LPS (1 microgram/ml), accumulated 1.4 and 3.5 nmol/10(6) cells nitrite, respectively. Addition of 100 microM L-arginine increased nitrite accumulation by 70 and 400% in control and lipopolysaccharide-treated AMPhi, respectively. Nitrite accumulation in the presence of L-arginine was reduced by poly-L-arginine and poly-L-lysine (100 and 300 microgram/ml) by 60 to 85% and 20 to 30%, respectively. Poly-L-arginine, but not poly-L-lysine, inhibited nitrite accumulation already in the absence of extracellular L-arginine. Poly-L-arginine (300 microgram/ml) inhibited [(3)H]-L-citrulline formation by AMPhi stronger than that of [(3)H]-L-ornithine. We conclude that cationic proteins can inhibit cellular transport of L-arginine and this can limit NO synthesis. Poly-L-arginine inhibits L-arginine uptake more effectively than other cationic proteins and exerts additional direct inhibitory effects on NO synthesis.
嗜酸性粒细胞衍生的阳离子蛋白在支气管哮喘的发病机制中起重要作用。我们测试了阳离子蛋白是否会干扰肺泡巨噬细胞（AMPhi）和气管上皮细胞中的阳离子氨基酸转运，以及L-精氨酸依赖性途径是否受到影响。研究了阳离子多肽对[(3)H]-L-精氨酸细胞摄取、亚硝酸盐积累以及一氧化氮（NO）合酶和精氨酸酶对[(3)H]-L-精氨酸的周转（分别形成[(3)H]-L-瓜氨酸和[(3)H]-L-鸟氨酸）的影响。聚-L-精氨酸以浓度依赖性方式降低大鼠AMPhi和气管上皮细胞中[(3)H]-L-精氨酸的摄取（在300微克/毫升时降低70%）。在大鼠AMPhi中测试的聚-L-赖氨酸、鱼精蛋白和主要碱性蛋白（每种浓度高达300微克/毫升）抑制[(3)H]-L-精氨酸摄取35%至50%。在无氨基酸的Krebs溶液中孵育6小时期间，在不存在或存在LPS（1微克/毫升）的情况下预培养的大鼠AMPhi分别积累1.4和3.5纳摩尔/10(6)个细胞的亚硝酸盐。添加100微摩尔/升L-精氨酸分别使对照和脂多糖处理的AMPhi中的亚硝酸盐积累增加70%和400%。在L-精氨酸存在下，聚-L-精氨酸和聚-L-赖氨酸（分别为100和300微克/毫升）使亚硝酸盐积累分别减少60%至85%和20%至30%。聚-L-精氨酸而非聚-L-赖氨酸在不存在细胞外L-精氨酸的情况下就抑制亚硝酸盐积累。聚-L-精氨酸（300微克/毫升）对AMPhi中[(3)H]-L-瓜氨酸形成的抑制作用强于[(3)H]-L-鸟氨酸。我们得出结论，阳离子蛋白可抑制L-精氨酸的细胞转运，这可能会限制NO的合成。聚-L-精氨酸比其他阳离子蛋白更有效地抑制L-精氨酸摄取，并对NO合成产生额外的直接抑制作用。
