Regulation of nitric oxide production in snail (Lymnaea stagnalis) defence cells:: a role for PKC and ERK signalling pathways
Regulation of nitric oxide production in snail (Lymnaea stagnalis) defence cells:: a role for PKC and ERK signalling pathways
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DOI：
10.1042/bc20050066
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发表时间：
2006-05-01
影响因子：
2.7
通讯作者：
Walker, Anthony J.
中科院分区：
文献类型：
--
作者：
Wright, Bernice;Lacchini, Audrey H.;Walker, Anthony J.
背景资料。一氧化氮（NO）是天然免疫反应中的重要分子。在软体动物中，NO由称为血细胞的移动的防御细胞产生;然而，调节这些细胞中NO产生的分子机制知之甚少。本研究的重点是细胞信号通路的作用，在NO生产的初级血细胞从蜗牛滞流。当用PMA（10 M）或β-1，3-葡聚糖海带多糖（10 mg/ml）攻击血细胞时，在60 min后分别观察到NO产生的8倍和4倍增加。NO合成酶抑制剂L-NAME（N-G-nitro-L-arginine methyl ester）和L-NMMA（N-G-monomethyl-L-arginine）可阻断海带多糖和PMA诱导的NO合成。用PMA或海带多糖处理血细胞也增加了PKC（蛋白激酶C）的磷酸化（激活）状态。当血细胞与PKC抑制剂（calphostin C或GF 109203 X）或ERK（细胞外信号调节激酶）通路抑制剂（PD 98059或U 0126）预孵育前的挑战，显着减少PKC和ERK磷酸化和NO的生产后，暴露于海带多糖或PMA观察。GF 109203 X和U 0126对NO产生的影响最大，PMA诱导的NO产生分别抑制了94%和87%，海带多糖诱导的NO产生分别抑制了50%和91%。这些数据表明，ERK和PKC组成的信号转导机制，调节NOS激活和随后的生产NO在软体动物血细胞的一部分。据我们所知，这是第一份报告，显示了这些信号蛋白在无脊椎动物防御细胞中产生NO的作用。
Background information. Nitric oxide (NO) is an important molecule in innate immune responses. In molluscs NO is produced by mobile defence cells called haemocytes; however, the molecular mechanisms that regulate NO production in these cells is poorly understood. The present study focused on the role of cell signalling pathways in NO production by primary haemocytes from the snail Lymnaea stagnalis.Results. When haemocytes were challenged with PMA (10 M) or the beta-1,3-glucan laminarin (10 mg/ml), an 8-fold and 4-fold increase in NO production were observed after 60 min respectively. Moreover, the NOS (NO synthase) inhibitors L-NAME (N-G-nitro-L-arginine methyl ester) and L-NMMA (N-G-monomethyl-L-arginine) were found to block laminarin- and PMA-induced NO synthesis. Treatment of haemocytes with PMA or laminarin also increased the phosphorylation (activation) status of PKC (protein kinase C). When haemocytes were preincubated with PKC inhibitors (calphostin C or GF109203X) or inhibitors of the ERK (extracellular-signal-regulated kinase) pathway (PD98059 or U0126) prior to challenge, significant reductions in PKC and ERK phosphorylation and NO production were observed following exposure to laminarin or PMA. The greatest effect on NO production was seen with GF109203X and U0126, with PMA-induced NO production inhibited by 94% and 87% and laminarin-induced NO production by 50% and 91% respectively.Conclusions. These data suggest that ERK and PKC comprise part of the signalling machinery that regulates NOS activation and subsequent production of NO in molluscan haemocytes. To our knowledge, this is the first report that shows a role for these signalling proteins in the generation of NO in invertebrate defence cells.