Department of Pharmaceutical Pathophysiology, Medical University of Gdansk, Gdansk, Poland.
Faculty of Chemistry, University of Warsaw, Warsaw, Poland.
Int J Nanomedicine. 2022 Sep 19;17:4383-4400. doi: 10.2147/IJN.S373691. eCollection 2022.
In the search for new drug delivery platforms for cardiovascular diseases and coating of medical devices, we synthesized eptifibatide-functionalized silver nanoparticles (AgNPs-EPI) and examined the pharmacological activity of AgNPs-EPI on platelets and endothelial cells in vitro and ex vivo.
Spherical AgNPs linked to eptifibatide were synthesized and characterized. Cytotoxicity was measured in microvascular endothelial cells (HMEC-1), platelets and red blood cells. Platelet mitochondrial respiration was measured using the Oxygraph-2k, a high-resolution modular respirometry system. The effect of AgNPs-EPI on the aggregation of washed platelets was measured by light aggregometry and the ex vivo occlusion time was determined using a reference laboratory method. The surface amount of platelet receptors such as P-selectin and GPIIb/IIIa was measured. The influence of AgNPS-EPI on blood coagulation science was assessed. Finally, the effect of AgNPs-EPI on endothelial cells was measured by the levels of 6-keto-PGF1alpha, tPa, cGMP and vWF.
We describe the synthesis of AgNPs using eptifibatide as the stabilizing ligand. The molecules of this drug are directly bonded to the surface of the nanoparticles. The synthesized AgNPs-EPI did not affect the viability of platelets, endothelial cells and erythrocytes. Preincubation of platelets with AgNPs-EPI protected by mitochondrial oxidative phosphorylation capacity. AgNPs-EPI inhibited aggregation-induced P-selectin expression and GPIIb/IIIa conformational changes in platelets. AgNPs-EPI caused prolongation of the occlusion time in the presence of collagen/ADP and collagen/adrenaline. AgNPs-EPI regulated levels of 6-keto-PGF1alpha, tPa, vWf and cGMP produced in thrombin stimulated HMEC-1 cells.
AgNPs-EPI show anti-aggregatory activity at concentrations lower than those required by the free drug acting via regulation of platelet aggregation, blood coagulation, and endothelial cell activity. Our results provide proof-of-principle evidence that AgNPs may be used as an effective delivery platform for antiplatelet drugs.
在寻找心血管疾病的新药物输送平台和医疗器械涂层的过程中，我们合成了依替巴肽功能化的银纳米粒子（AgNPs-EPI），并在体外和体内研究了 AgNPs-EPI 对血小板和内皮细胞的药理学活性。
合成并表征了与依替巴肽连接的球形 AgNPs。在微血管内皮细胞（HMEC-1）、血小板和红细胞中测量细胞毒性。使用高分辨率模块化呼吸测定系统 Oxygraph-2k 测量血小板线粒体呼吸。通过光聚合度法测量 AgNPs-EPI 对洗涤血小板聚集的影响，并使用参考实验室方法确定体外闭塞时间。测量血小板受体如 P-选择素和 GPIIb/IIIa 的表面量。评估 AgNPS-EPI 对血液凝固科学的影响。最后，通过 6-酮-PGF1alpha、tPA、cGMP 和 vWF 的水平测量 AgNPs-EPI 对内皮细胞的影响。
我们描述了使用依替巴肽作为稳定配体合成 AgNPs 的方法。该药物的分子直接键合到纳米粒子的表面。合成的 AgNPs-EPI 不影响血小板、内皮细胞和红细胞的活力。AgNPs-EPI 预处理可保护血小板线粒体氧化磷酸化能力。AgNPs-EPI 抑制了血小板聚集诱导的 P-选择素表达和 GPIIb/IIIa 构象变化。在胶原/ADP 和胶原/肾上腺素存在下，AgNPs-EPI 导致闭塞时间延长。AgNPs-EPI 调节了凝血酶刺激的 HMEC-1 细胞中产生的 6-酮-PGF1alpha、tPA、vWf 和 cGMP 的水平。
AgNPs-EPI 在低于游离药物所需浓度下表现出抗聚集活性，通过调节血小板聚集、血液凝固和内皮细胞活性发挥作用。我们的结果提供了初步证据，证明 AgNPs 可用作抗血小板药物的有效输送平台。
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