二甲亚砜与海藻糖联用对冷冻保存血小板的保护作用

Protection of Cryopreserved Platelets by Dimethyl Sulfoxide Combined with Trehalose

本研究观察二甲亚砜(DMSO)与海藻糖联用对冷冻保存血小板的保护效果。实验分设空白组、海藻糖组、DMSO组、5%DMSO加海藻糖联用组、2.5%DMSO加海藻糖联用组。各组血小板置-80℃冰箱保存,37℃水浴融化。采用血细胞计数仪测定血小板回收率和MPV值、电子显微镜观察血小板超微结构变化和流式细胞仪测定血小板的CD41、CD42b、CD61及CD62p的表达水平。结果表明:海藻糖单独应用对提高回收率的保护作用不强,但海藻糖处理后冷冻保存的血小板的形态接近正常。DMSO在保证冷冻保存血小板的回收率和血小板整体完好性方面作用较为突出,但其血小板形态偏向于肿胀,仍有部分血小板呈异形性改变。DMSO和海藻糖合用对维持血小板外部形态和内部结构接近正常稳态方面具有保护作用,同时保证冷冻保存血小板具有理想的回收率和较高的CD41、CD42b、CD61、CD62p表达水平。结论:DMSO和海藻糖联合应用对冷冻保存血小板具有协同保护作用,但DMSO和海藻糖单用或合用都较难抑制冷冻保存血小板的活化,两者合用作为血小板冷冻保护剂有望促使冷冻保存血小板临床输注效果的进一步提高。

This study was aimed to investigate the protective effects of dimethylsulfoxide （DMSO） combined with trehalose on the cryopreserved platelets. The platelets were preserved at - 80℃. The experiments were divided into 5 groups： blank control group composed of apheresis platelet suspension; trehalose group composed of apheresis plalelet suspension and 0.25 mol/L trehalose; DMSO group composed of apheresis platelet suspension and 5% DMSO; 5% combined group composed of apheresis platelet suspension, 5% DMSO and 0.25 mot/L trehalose; 2.5% combined group composed of apheresis platelet suspension, 2.5% DMSO and 0.25 mol/L threhalose. All the groups were thawed at 37℃ in a waterbath. The recovery rate of platelets and mean platelet volume （MPV） were assayed by using hemocytometer; the ultrastructural changes were examined by electron microscopy; the expressions of CD41, CD42b, CD61 and CD62p on platelets were detected by flow cytometry. The results indicated that single use of trehalose had no strong effect in increasing the recovery rate of platelets, but the morphology of platelets was close to normal. The DMSO showed significant effect in increasing the recovery rate of platelets and maintaining the intact property of platelets, however, the shape of platelets tended to seelling, and partial platelets still displayed heteromorphic changes. The combination of DMSO and trehalose revealed the protective effect on the external morphology and internal structure of platelets to be close to the normal homeostasis, and ensured an ideal recovery rate of the cryopreserved platelets and higher expression levels of CD41, CD42b, CD61 and CD62p in the same time. It is concluded that the combined use of DMSO and trehalose possesses the synergistic protective effect on the cryopreserved platelets, therefore, the combined use of both as the protective agent is hopeful to further raise the effectiveness of clinical infusion of the cryopreserved platelets.