Almeida Cardelli Nádia J, Elisa Lopes-Pires M, Bonfitto Pedro H L, Ferreira Heloisa H, Antunes Edson, Marcondes Sisi
Department of Pharmacology, Faculty of Medical Sciences, University of Campinas (UNICAMP), Campinas, SP, Brazil.
Laboratory of Inflammation Research, São Leopoldo Mandic Institute and Research Center, Campinas, Sao Paulo, Brazil.
Life Sci. 2017 Mar 1;172:2-7. doi: 10.1016/j.lfs.2016.12.013. Epub 2016 Dec 23.
Cross-talk between platelets and lymphocytes may play a role in different pathological conditions like sepsis. This study aimed to investigate the effect of lymphocytes on platelet aggregation in lipopolysaccharide (LPS)-stimulated and non-stimulated cells.
Lymphocytes and platelet-rich plasma (PRP) were obtained from rat arterial blood. Platelets (1.2×10platelets/ml) were incubated with lymphocytes (0.8×10cells/ml) in the presence or not of LPS (100μg/ml), after which ADP (5μM)-induced platelet aggregation was carried out.
Lymphocytes inhibited by 51% the platelet aggregation, which was significantly prevented by the non-selective NO inhibitor l-NAME (300μM) or the selective iNOS inhibitor 1400W (100μM), as well as by the soluble guanylyl cyclase (sGC) inhibitor ODQ (10μM). The platelet inhibition by lymphocytes was accompanied by 2-fold increase of intraplatelet cGMP levels. Next, lymphocytes and platelets were co-incubated with LPS for 6h. In LPS-treated cells, lymphocytes produced a larger inhibition of platelet aggregation (62%), despite the same elevation of cGMP levels (2.2-fold increase). This inhibitory effect was prevented by l-NAME and 1400W, but rather unaffected by ODQ. The peroxynitrite (ONOO) scavenger -(-)epigallocatechin gallate (ECG, 100μM) abolished the inhibition by lymphocytes on platelet aggregation in LPS-treated cells, but not in non-treated cells.
Our results show that lymphocytes act to inhibit platelet aggregation via iNOS-derived NO release and cGMP generation. In presence of LPS, ONOO production accounts for the platelet inhibition.
血小板与淋巴细胞之间的相互作用可能在脓毒症等不同病理状态中发挥作用。本研究旨在探究淋巴细胞对脂多糖（LPS）刺激及未刺激细胞中血小板聚集的影响。
从大鼠动脉血中获取淋巴细胞和富血小板血浆（PRP）。将血小板（1.2×10⁹个血小板/毫升）与淋巴细胞（0.8×10⁶个细胞/毫升）在有或无LPS（100微克/毫升）的情况下孵育，之后进行ADP（5微摩尔）诱导的血小板聚集实验。
淋巴细胞使血小板聚集受到51%的抑制，非选择性NO抑制剂L-NAME（300微摩尔）、选择性诱导型一氧化氮合酶（iNOS）抑制剂1400W（100微摩尔）以及可溶性鸟苷酸环化酶（sGC）抑制剂ODQ（10微摩尔）可显著阻止这种抑制作用。淋巴细胞对血小板的抑制作用伴随着血小板内cGMP水平升高两倍。接下来，将淋巴细胞和血小板与LPS共同孵育6小时。在LPS处理的细胞中，淋巴细胞对血小板聚集的抑制作用更大（62%），尽管cGMP水平升高幅度相同（增加2.2倍）。这种抑制作用可被L-NAME和1400W阻止，但不受ODQ影响。过氧亚硝酸盐（ONOO）清除剂（-）表没食子儿茶素没食子酸酯（ECG，100微摩尔）可消除淋巴细胞对LPS处理细胞中血小板聚集的抑制作用，但对未处理细胞无此作用。
我们的结果表明，淋巴细胞通过iNOS衍生的NO释放和cGMP生成来抑制血小板聚集。在存在LPS的情况下，ONOO的产生是导致血小板抑制的原因。
