ROLES FOR PROTEIN-KINASES IN THE INDUCTION OF NITRIC-OXIDE SYNTHASE IN ASTROCYTES
ROLES FOR PROTEIN-KINASES IN THE INDUCTION OF NITRIC-OXIDE SYNTHASE IN ASTROCYTES
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DOI：
10.1002/glia.440110303
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发表时间：
1994-07-01
期刊：
影响因子：
6.2
通讯作者：
MURPHY, S
中科院分区：
文献类型：
--
作者：
SIMMONS, ML;MURPHY, S
脂多糖（LPS）或干扰素（IFN）-γ和白细胞介素（IL）-1 β的组合可以在星形胶质细胞培养物中诱导钙非依赖性一氧化氮合酶（iNOS）（Simmons和Murphy：J Neurochem 59：897，1992; Eur J Neurosci 5：825，1993; Galea等：Proc Natl Acad Sci USA 89：10945，1992）。这种诱导可以通过测定培养物中的环GMP水平来测量，其与亚硝酸盐积累的测量相关，但比亚硝酸盐积累的测量更敏感。为了研究潜在的第二信使系统参与诱导的诱导型一氧化氮合酶，佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA），蛋白激酶C（PKC）激活剂，和各种蛋白激酶抑制剂。PMA可诱导cGMP浓度升高，呈时间、剂量和L-精氨酸依赖性，地塞米松或放线菌素D可抑制PMA诱导的cGMP浓度升高。这种诱导可以通过与IFN-γ同时治疗而显著增加。与特异性cDNA探针杂交可证明iNOS mRNA的存在。H7（一种非特异性丝氨酸/苏氨酸激酶抑制剂）而不是H89（一种更特异性的PKA抑制剂）阻止了所有试剂的诱导。然而，下调PKC或用PKC抑制剂calphostin C预处理并不能阻止LPS或细胞因子的诱导，这表明PKC对于这些介质诱导iNOS并不是必需的。此外，染料木黄酮（一种非特异性酪氨酸激酶抑制剂）可以防止所有代理人的诱导，但更具体的抑制剂，tyrphostin，衰减只有NOS诱导LPS。这些结果表明，PKC的激活可以导致，但不是必需的，诱导NOS在星形胶质细胞和有一个潜在的作用，酪氨酸激酶在NOS诱导LPS。这种复杂的控制诱导的iNOS，似乎需要激活多个途径的最大效果，可能是一种保障措施，以防止生产的潜在毒性一氧化氮（NO）在正常的生理条件下。(C)1994 Wiley-Liss，Inc.
Lipopolysaccharide (LPS) or a combination of interferon (IFN)-gamma and interleukin (IL)-1 beta can induce a calcium-independent nitric oxide synthase (iNOS) in astrocyte cultures (Simmons and Murphy: J Neurochem 59:897, 1992; Eur J Neurosci 5:825, 1993; Galea et al: Proc Natl Acad Sci USA 89:10945, 1992). This induction can be measured by assaying cyclic GMP levels in the cultures, which correlates with, but is more sensitive than, measurement of nitrite accumulation. To study potential second-messenger systems involved in the induction of iNOS, phorbol 12-myristate 13-acetate (PMA), a protein kinase C (PKC) activator, and various protein kinase inhibitors were employed. PMA induced a time-, dose-, and L-arginine-dependent increase in cyclic GMP, which could be inhibited by dexamethasone or actinomycin D. This induction could be dramatically increased by concurrent treatment with IFN-gamma. The presence of iNOS mRNA could be demonstrated by hybridization with a specific cDNA probe. H7 (a non-specific serine/threonine kinase inhibitor) but not H89 (a more specific PKA inhibitor) prevented induction by all agents. However, downregulation of PKC or pretreatment with the PKC inhibitor calphostin C did not prevent the induction by LPS or cytokines, suggesting that PKC is not necessary for iNOS induction by these mediators. Additionally, genistein (a nonspecific tyrosine kinase inhibitor) could prevent induction by all agents, but the more specific inhibitor, tyrphostin, attenuated only NOS induction by LPS. These results suggest that activation of PKC can lead to, but is not necessary for, the induction of NOS in astrocytes and that there is a potential role for tyrosine kinases in NOS induction by LPS. This complex control of induction of iNOS, seemingly requiring activation of multiple pathways for maximal effect, might represent a safeguard to prevent production of potentially toxic nitric oxide (NO) under normal physiological conditions. (C) 1994 Wiley-Liss, Inc.