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蛋白激酶c和蛋白酪氨酸激酶介导脂多糖及细胞因子诱导大鼠血管平滑肌细胞一氧化氮的生成

韩雅玲, 康建, 李少华

沈阳军区总医院全军心血管内外科研究所心内科，沈阳110016

摘要

采用Sprague- Dawley大鼠胸主动脉中膜、外膜和培养的血管平滑肌细胞(VS MCs)作材料，鉴定不同类型的血管组织经炎性介质刺激后其一氧化氮(NO)的产生来源，阐明蛋白激酶C (PKC)和蛋白酪氨酸激酶( PTK)介导大鼠US MCs生成NO的调控机制。大鼠US MCs经脂多糖(LPS)和细胞因子(TNF a , IL- 1 [3)处理后，以剂量依赖方式促进NO释放。采用stern Blot证实经刺激的US MCs伴有i NOS表达上调。进一步实验表明PKC和PTK参与LPS和细胞因子诱导NO生成的胞内信号转导。用P KC抑制剂H7与VS MCs共培育，H7能明显减少LPS . TNF。和IL-场诱导细胞NO的形成。白屈菜赤碱亦可抑制NO的生成，但HA1004对US MCs的NO生成无抑制作用，提示PKC参与NO的生成与调控。PTK抑制剂genistein和tyrphostin AG18均能抑制由LPS . TNF。和IL-1 [3引发US MCs释放NO，同时伴i NOS蛋白表达下调，而P KC抑制剂不能阻断i NOS的表达。上述观察结果提示，PKC介导LPS和细胞因子诱导细胞合成NO可能是通过i NOS翻译后加工;而PTK则以上调i NOS表达而促增NO生成。

关键词： 一氧化氮; 蛋白激酶C; 蛋白酪氨酸激酶; 平滑肌

Protein Kinase C and protein tyrosine Kinase mediate lipopolysaccharide- and cytokine-induced nitric oxide formationin vascular smooth muscle cells of rats

HAN Ya- Ling, KANG Jian, LI Shao Hua

Department of Cardiology，General Hospital of ShenyanA，The Institute of Cardiovascular Research，PLA, 110016 Department of Pathology and Laboratory Medicine，Robert Wood Johnson Medical School，New Jersey 08854，USA

Abstract

Rat aorta media，adventitia and cultured vascular smooth muscle cells(VSMCs) were used in this stndy to identfy the source of nitric oxide(NO) generation from various cell types of vascular tissues and to elucidate the mechanisms involved in NO formation .Treatment of vascular media and VSMCs with lipopolysaccharide(LPS) or cytokines[tumor necrosis factor alpha(TNF。)
and interleukirrl beta(IL- Ip)]resulted in a dose-dependent increase of NO release .Inducible nitric oxide synthase(iNOS) in the stimulated VSMCs was significantly upregulated as shown by Western blot analysis .Protein kinase C(PK()inhibitor 1一(5- isoquinolinesulfonyl)一2- methylpiperazine(117) prevented LPSL，TNF a- and IL 1 [r induce d NO production，whereas N(2- guanidinoethyl)一5-isoquinoline- sulfonamide(HA1004)，an H7 analogue with little activity towards PK(，had no inhibition effect .The role of PKC in
LPS- and cytokine-induced changes on NO formation was confirmed by using another structurally distinct PKC inhibitor chelerythrine.Treatment of VSMCs with protein tyrosine kinase(PT)nhibitor genistein or tyrphostin AG18 also reduced the NO production evoked by LPS，TNF a or IL 1 [3，which was ssociated with inhibition of iNO8 protein expression .In contrast，PKC inhibitorythrine did not affect iNOS expression .These results suggest that PTK mediates LPS- and cytokine-induced NO formation by upregulation of iNO8 expression .PKC may be involved in the post translational modification of iNO8 or the regulation of the availability of iNOS substrates and cofactors.

Key words: Nitric oxide; nrntein kinase C;nrntein tvrmine kinase ; smooth muscle

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引用本文：

韩雅玲, 康建, 李少华. 蛋白激酶c和蛋白酪氨酸激酶介导脂多糖及细胞因子诱导大鼠血管平滑肌细胞一氧化氮的生成[J]. 生理学报 2003; 55 (3): .

HAN Ya- Ling, KANG Jian, LI Shao Hua. Protein Kinase C and protein tyrosine Kinase mediate lipopolysaccharide- and cytokine-induced nitric oxide formationin vascular smooth muscle cells of rats . Acta Physiol Sin 2003; 55 (3): (in Chinese with English abstract).