Bertino A M, Qi X Q, Li J, Xia Y, Kuter D J
Department of Medicine, Division of Hematology and Oncology, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, USA.
Transfusion. 2003 Jul;43(7):857-66. doi: 10.1046/j.1537-2995.2003.t01-4-00431.x.
PLTs for transfusion lose viability during storage in blood banking. This loss of viability is accelerated at 37 degrees C, as is the risk of bacterial contamination, and has led to the selection of 22 degrees C as the routine storage temperature. Because PLTs contain an intact apoptotic mechanism, we sought to determine whether PLTs undergo apoptosis during storage and whether storage at 37 degrees C accelerated this process.
PLT-rich plasma from PLT concentrates was stored at 37 or 22 degrees C in small aliquots or whole bags, with and without cell-permeable caspase inhibitors. Number of PLTs, pH, LDH level, and 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium activity were analyzed over time. PLT lysates were prepared and tested for the presence and activation of apoptotic proteins by enzyme assay and Western blotting.
PLT viability was greatly reduced after 1 to 2 days of storage at 37 degrees C; however, signs of apoptosis were evident by 3 hours after temperature shift. In temperature-stressed PLTs only, a gradual rise in caspase-3 activity was detected that correlated with the appearance of the 17- to 20-kDa cleavage products of caspase-3. Gelsolin, a caspase-3 substrate, underwent cleavage within the same time frame. Bcl-xL and caspase-2 also declined significantly; caspase-9 activity rose. Specific caspase inhibitors could prevent caspase activation but did not improve PLT cellular viability at 37 degrees C.
PLTs contain apoptotic proteins that are activated during PLT storage at 37 degrees C and may account for the rapid decline in PLT cellular viability. Although ineffective here, inhibition of PLT apoptosis may improve PLT cellular viability.
用于输血的血小板在血库储存期间会丧失活力。这种活力丧失在37℃时会加速，细菌污染风险也会增加，这促使人们选择22℃作为常规储存温度。由于血小板含有完整的凋亡机制，我们试图确定血小板在储存期间是否会发生凋亡，以及在37℃储存是否会加速这一过程。
将血小板浓缩物中的富血小板血浆以小份或整袋形式分别在37℃或22℃下储存，同时添加或不添加细胞可渗透的半胱天冬酶抑制剂。随着时间的推移，分析血小板数量、pH值、乳酸脱氢酶水平和3-（4,5-二甲基噻唑-2-基）-5-（3-羧甲氧基苯基）-2-（4-磺基苯基）-2H-四唑活性。制备血小板裂解物，并通过酶测定和蛋白质印迹法检测凋亡蛋白的存在和激活情况。
在37℃储存1至2天后，血小板活力大幅降低；然而，在温度转变后3小时，凋亡迹象就很明显。仅在温度应激的血小板中，检测到半胱天冬酶-3活性逐渐升高，这与半胱天冬酶-3 17至20 kDa裂解产物的出现相关。凝溶胶蛋白作为半胱天冬酶-3的底物，在同一时间框架内发生裂解。Bcl-xL和半胱天冬酶-2也显著下降；半胱天冬酶-9活性升高。特异性半胱天冬酶抑制剂可阻止半胱天冬酶激活，但在37℃时并不能提高血小板细胞活力。
血小板含有在37℃储存期间被激活的凋亡蛋白，这可能是血小板细胞活力迅速下降的原因。虽然在此处无效，但抑制血小板凋亡可能会提高血小板细胞活力。
