Institute of Transfusion Medicine and Haemotherapy, University of Wuerzburg, Oberduerrbacher Straße 6, D-97080 Wuerzburg, Germany.
Cell Signal. 2024 Oct;122:111306. doi: 10.1016/j.cellsig.2024.111306. Epub 2024 Jul 22.
Platelet concentrates (PC) are used to treat patients with thrombocytopenia and hemorrhage, but there is still the demand to find the optimal strategy for temperature-dependent storage of PC. Recently, we could show that cold storage for 1 h (short-term refrigeration) is sufficient to induce enhanced platelet responsiveness. The aim of this study was to investigate effects of cold storage on collagen-dependent activating signalling pathways in platelets from apheresis-derived PC (APC). APC on day 1 or day 2 of storage, were either continuously kept at room temperature (RT, 22 °C), or for comparison, additionally kept at cold temperature (CT, 4 °C) for 1 h. CD62P expression was determined by flow cytometry. Western Blot technique was used to analyze collagen-induced phosphorylation of p38, ERK1/2 or Akt/PKB and its inhibition by prostaglandin E1 (PGE1) or nitric monoxide donor. Adhesion of platelets on collagen-coated surfaces and intracellular phosphorylation of vasodilator-stimulated phosphoprotein (VASP) was visualized by immune fluorescence microscopy. CD62P expression was increased after short-term refrigeration. CT exposition for 1 h induced an elevation of basal ERK1/2 phosphorylation and an alleviation of PGE1- or DEA/NO-suppressed ERK1/2 phosphorylation in APC on day 1 and 2 of storage. Similar, but more moderate effects were observable for p38 phosphorylation. Akt/PKB phosphorylation was increased only in APC on day 2. Refrigeration for 1 h promoted platelet adhesion and reduced basal VASP phosphorylation in adherent platelets. The attenuation of inhibitory signalling in short-term refrigerated stored platelets is associated with enhanced reactivity of activating signalling pathways, especially ERK1/2. Functionally, these processes correlate with increased adhesion of refrigerated platelets on collagen-coated surfaces. The results help to further optimize temperature-dependent strategies for platelet storage.
血小板浓缩物 (PC) 用于治疗血小板减少症和出血患者，但仍需要寻找最佳策略来实现 PC 的温度依赖性储存。最近，我们发现冷藏 1 小时（短期冷藏）足以诱导增强的血小板反应性。本研究的目的是研究冷藏对来自单采 PC (APC) 的胶原依赖性激活信号通路的影响。第 1 天或第 2 天储存的 APC 要么连续保持室温（22°C，RT），要么与室温相比，另外冷藏 1 小时（4°C，CT）。通过流式细胞术测定 CD62P 表达。采用 Western Blot 技术分析胶原诱导的 p38、ERK1/2 或 Akt/PKB 磷酸化及其对前列腺素 E1 (PGE1) 或一氧化氮供体的抑制作用。通过免疫荧光显微镜观察血小板在胶原涂层表面的黏附和血管扩张刺激磷蛋白 (VASP) 的细胞内磷酸化。短期冷藏后 CD62P 表达增加。CT 暴露 1 小时可诱导储存第 1 天和第 2 天的 APC 中基础 ERK1/2 磷酸化升高，并减轻 PGE1 或 DEA/NO 抑制的 ERK1/2 磷酸化。p38 磷酸化也可观察到类似但更为温和的作用。仅在储存第 2 天的 APC 中 Akt/PKB 磷酸化增加。冷藏 1 小时可促进血小板黏附并降低黏附血小板中的基础 VASP 磷酸化。短期冷藏储存的血小板中抑制信号的衰减与激活信号通路，尤其是 ERK1/2 的反应性增强有关。功能上，这些过程与冷藏血小板在胶原涂层表面的黏附增加有关。这些结果有助于进一步优化血小板储存的温度依赖性策略。
