Ozawa Naoko, Shichiri Masayoshi, Iwashina Masatora, Fukai Nozomi, Yoshimoto Takanobu, Hirata Yukio
Department of Clinical and Molecular Endocrinology, Tokyo Medical and Dental University Graduate School, Tokyo, Japan.
Hypertens Res. 2004 Feb;27(2):93-9. doi: 10.1291/hypres.27.93.
Shear stress caused by blood flow is a potent physiological stimulus for the generation of nitric oxide (NO) in endothelial cells, which is believed to derive from the up-regulation and post-transcriptional activation of endothelial constitutive NO synthase (ecNOS). However, it has yet to be demonstrated that inducible NO synthase (iNOS) plays a significant role in shear stress-induced NO production from endothelial cells. We used parallel plate-type flow chambers that detect fluid shear stress to determine that shear stress, as quantified by a real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR), increased iNOS gene transcripts in cultured endothelial cells, which resulted in increased NO production. Shear stress-induced iNOS expression was inhibited by pyrrolidine dithiocarbamate (PDTC), an antioxidant and nuclear factor kappaB (NF-kappaB) blocker, and by MG132, an aldehyde peptide proteasome inhibitor that antagonizes I kappaB-kinase. Laminar shear stress increased the transcriptional activity of NF-kappaB, whereas over-expression of an I kappaB-alpha mutant that inhibits the activation of NF-KB in a dominant-negative fashion was found to attenuate the induction of endothelial iNOS by shear stress. The present results demonstrate that shear stress induces iNOS in the endothelium, mainly via the activation of NF-kappaB.
血流引起的剪切应力是内皮细胞中一氧化氮（NO）生成的一种强大生理刺激，据信这源于内皮型组成型一氧化氮合酶（ecNOS）的上调和转录后激活。然而，诱导型一氧化氮合酶（iNOS）在内皮细胞剪切应力诱导的NO产生中是否起重要作用尚未得到证实。我们使用能检测流体剪切应力的平行板型流动腔来确定，通过实时定量逆转录聚合酶链反应（RT-PCR）量化的剪切应力会增加培养的内皮细胞中iNOS基因转录本，进而导致NO产生增加。吡咯烷二硫代氨基甲酸盐（PDTC），一种抗氧化剂和核因子κB（NF-κB）阻滞剂，以及MG132，一种拮抗IκB激酶的醛肽蛋白酶体抑制剂，均可抑制剪切应力诱导的iNOS表达。层流剪切应力增加了NF-κB的转录活性，而以显性负性方式抑制NF-κB激活的IκB-α突变体的过表达被发现可减弱剪切应力对内皮iNOS的诱导。目前的结果表明，剪切应力主要通过激活NF-κB在内皮中诱导iNOS。
