Chang Yi, Hsu Wen-Hsien, Lu Wan-Jung, Jayakumar Thanasekaran, Liao Jiun-Cheng, Lin Mei-Jiun, Wang Shwu-Huey, Geraldine Pitchairaj, Lin Kuan-Hung, Sheu Joen-Rong
250 Wu-Hsing St., Graduate Institute of Medical Sciences, Taipei Medical University, Taipei 110, Taiwan.
Curr Pharm Biotechnol. 2015;16(5):451-61.
CME-1 is a polysaccharide purified from the mycelia of medicinal mushroom Cordyceps sinensis, its molecular weight was determined to be 27.6 kDa by using nuclear magnetic resonance and gas chromatography-mass spectrometry. The initiation of arterial thromboses is relevant to various cardiovascular diseases (CVDs) and is believed to involve platelet activation. Our recent study exhibited that CME-1 has potent antiplatelet activity via the activation of adenylate cyclase/cyclic AMP ex vivo and in vivo.
The aggregometry, and immunoblotting were used in this study.
In this study, the mechanisms of CME-1 in platelet activation is further investigated and found that CME-1 inhibited platelet aggregation as well as the ATP-release reaction, relative intracellular [Ca(+2)] mobilization, and the phosphorylation of phospholipase C (PLC)γ2 and protein kinase C (PKC) stimulated by collagen. CME-1 has no effects on inhibiting either convulxin, an agonist of glycoprotein VI, or aggretin, an agonist of integrin α2β1 stimulated platelet aggregation. Moreover, this compound markedly diminished thrombin and arachidonic acid (AA) induced phosphorylation of p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinase 2, c-Jun N-terminal kinase 1, and Akt. Treatment with SQ22536, an inhibitor of adenylate cyclase, markedly diminished the CME-1-mediated increasing of cyclic AMP level and reversed prostaglandin E1- or CME-1-mediated inhibition of platelet aggregation and p38 MAPK and Akt phosphorylation stimulated by thrombin or AA. Furthermore, phosphodiesterase activity of human platelets was not altered by CME-1.
The crucial finding of this study is that the antiplatelet activity of CME-1 may initially inhibit the PLCγ2-PKC-p47 cascade, and inhibit PI3-kinase/Akt and MAPK phosphorylation through adenylate cyclase/ cyclic AMP activation, then inhibit intracellular [Ca(+2)] mobilization, and, ultimately, inhibit platelet activation. The novel role of CME-1 in antiplatelet activity indicates that this compound exhibits high therapeutic potential for treating or preventing CVDs.
CME - 1是从药用真菌冬虫夏草菌丝体中纯化得到的一种多糖，通过核磁共振和气相色谱 - 质谱联用测定其分子量为27.6 kDa。动脉血栓形成与多种心血管疾病（CVD）相关，并且被认为涉及血小板活化。我们最近的研究表明，CME - 1在体内外均可通过激活腺苷酸环化酶/环磷酸腺苷发挥强大的抗血小板活性。
本研究采用了血小板聚集测定法和免疫印迹法。
在本研究中，进一步探究了CME - 1在血小板活化中的作用机制，发现CME - 1可抑制血小板聚集以及ATP释放反应、细胞内相对钙离子动员，以及由胶原蛋白刺激的磷脂酶C（PLC）γ2和蛋白激酶C（PKC）的磷酸化。CME - 1对抑制糖蛋白VI激动剂convulxin或整合素α2β1激动剂aggretin刺激的血小板聚集均无作用。此外，该化合物显著降低了凝血酶和花生四烯酸（AA）诱导的p38丝裂原活化蛋白激酶（MAPK）、细胞外信号调节激酶2、c - Jun氨基末端激酶1和Akt的磷酸化。用腺苷酸环化酶抑制剂SQ22536处理，显著降低了CME - 1介导的环磷酸腺苷水平升高，并逆转了前列腺素E1或CME - 1介导的对血小板聚集的抑制以及凝血酶或AA刺激的p38 MAPK和Akt磷酸化。此外，CME - 1未改变人血小板的磷酸二酯酶活性。
本研究的关键发现是，CME - 1的抗血小板活性可能首先抑制PLCγ2 - PKC - p47级联反应，并通过腺苷酸环化酶/环磷酸腺苷激活抑制PI3激酶/Akt和MAPK磷酸化，进而抑制细胞内钙离子动员，最终抑制血小板活化。CME - 1在抗血小板活性方面的新作用表明，该化合物在治疗或预防心血管疾病方面具有很高的治疗潜力。
