Yemisci Muge, Ay Hakan, Kocaefe Cetin, Qui Jianhua, Topalkara Kamil, Ozgüç Meral, Kirazli Serafettin, Ozcebe Osman, Moskowitz Michael A, Dalkara Turgay
Department of Neurology, Faculty of Medicine, Hacettepe University, Ankara, Turkey.
Cerebrovasc Dis. 2008;26(2):190-8. doi: 10.1159/000145327. Epub 2008 Jul 15.
BACKGROUND/AIMS: Experimental studies suggest an enhanced endothelial and platelet nitric oxide (NO) generation after statin treatment, possibly due to increased endothelial NO synthase (eNOS) activity and protein levels. In parallel with experimental research, statins were shown to increase the forearm blood flow independently of serum cholesterol in humans. However, it was not possible to correlate blood flow changes with eNOS levels in these studies due to limitations in obtaining arterial samples. Hence, we investigated changes in eNOS activity, mRNA and protein levels after statin treatment in human platelets, which are readily accessible unlike arteries.
In vitro bleeding times were measured in 22 patients by stimulating platelets with collagen-epinephrine or collagen-ADP. To assess platelet eNOS activity, the bleeding times were also determined after incubating platelets with L-arginine. The measurements were repeated following 14 days of pravastatin (40 mg/day) treatment. Platelet-rich plasma was collected before and after statin treatment to evaluate eNOS mRNA (semiquantitative RT-PCR) and protein levels (Western blotting).
The basal bleeding time was prolonged by 24 +/- 3% (mean +/- SE) when the samples were incubated with 500 microM of L-arginine. The NOS inhibitor L-N(5)-(I-iminoethyl)ornithine reversed this effect, suggesting that it was mediated by NO. After statin treatment, the NO-mediated prolongation of the bleeding time with 500 microM of L-arginine was significantly potentiated (to 44 +/- 10%). Despite enhanced eNOS activity, there was no significant change in platelet eNOS mRNA and protein levels after statin treatment.
These data demonstrate that platelet eNOS activity is potentiated after statin treatment in humans in parallel with experimental studies.
背景/目的：实验研究表明，他汀类药物治疗后内皮细胞和血小板一氧化氮（NO）生成增加，这可能是由于内皮型一氧化氮合酶（eNOS）活性和蛋白水平升高所致。与实验研究同步，在人体中发现他汀类药物可独立于血清胆固醇增加前臂血流量。然而，由于获取动脉样本存在局限性，在这些研究中无法将血流变化与eNOS水平相关联。因此，我们研究了他汀类药物治疗后人血小板中eNOS活性、mRNA和蛋白水平的变化，因为血小板与动脉不同，易于获取。
通过用胶原-肾上腺素或胶原-ADP刺激血小板，测量22例患者的体外出血时间。为评估血小板eNOS活性，在用L-精氨酸孵育血小板后也测定出血时间。在普伐他汀（40mg/天）治疗14天后重复测量。在他汀类药物治疗前后收集富含血小板的血浆，以评估eNOS mRNA（半定量RT-PCR）和蛋白水平（蛋白质印迹法）。
当样本与500μM L-精氨酸孵育时，基础出血时间延长了24±3%（平均值±标准误）。一氧化氮合酶抑制剂L-N(5)-(I-亚氨乙基)鸟氨酸可逆转这种效应，表明其由NO介导。他汀类药物治疗后，500μM L-精氨酸介导的出血时间延长显著增强（至44±10%）。尽管eNOS活性增强，但他汀类药物治疗后血小板eNOS mRNA和蛋白水平无显著变化。
这些数据表明，与实验研究一致，他汀类药物治疗后人血小板中的eNOS活性增强。
