a Ænitis technologies S.A.S , Hôpital Saint-Louis , Paris , France.
b INSERM U1148 , Hôpital Bichat , Paris , France.
Platelets. 2019;30(2):174-180. doi: 10.1080/09537104.2017.1386296. Epub 2017 Dec 6.
Purity, limited platelet activation, and preservation of platelet function are important stakes of preparation of platelet concentrates (PC) for clinical use. In fact, contaminating red blood cells and leukocytes, as well as activated and/or poorly functional platelets in PC, represents a risk of poor efficiency and adverse side effects during platelet transfusion. Therefore, optimization of preparation and storage of PC is still an active field of research. Shear-induced platelet activation is an unwanted side effect of the hard-spin (up to 5000g) step of centrifugation-based methods currently used in blood banks to prepare PC from whole blood samples. Here, we evaluated the effectiveness of an acoustic-based fractionation device for the isolation of human platelets from whole blood bags. The purity, activation status, and functionality of platelets isolated by acoustopheresis were compared with those of platelets isolated using a reference protocol known to produce limited platelet activation and consisting of two consecutive soft-spin centrifugations (120g and 1200g). Platelet concentration and purity were determined using an automated hematology analyzer. Platelet activation status and platelet reactivity to collagen and thrombin were assessed in flow cytometry by measurement of surface expression of P-selectin and activated integrin αIIbβ3. The ability of isolated platelets to incorporate into a thrombus when transfused to NOD/SCID mice was investigated by intravital microscopy using the ferric chloride-induced thrombosis model. Blood fractionation by acoustophoresis led to the elimination of more than 80% of red blood cells and leukocytes from the platelet fraction, whose mean purity was of 92.8 ± 12.8%. The activation status and reactivity to collagen and thrombin of acoustophoresis-isolated platelets were similar to those of platelets isolated by soft-spin centrifugation. Finally, acoustophoresis-isolated platelets were tethered, adhered to the vessel wall, and incorporated into a growing thrombus following ferric chloride-induced vascular injury. Together, our results indicate that acoustophoresis is a suitable method for the isolation of human platelets with minimal platelet activation and preservation of platelet function.
纯度、有限的血小板激活和血小板功能的保存是临床应用血小板浓缩物（PC）制备的重要关注点。事实上，PC 中污染的红细胞和白细胞以及激活和/或功能不良的血小板，代表着血小板输注过程中效率低下和不良反应的风险。因此，优化 PC 的制备和储存仍然是一个活跃的研究领域。剪切诱导的血小板激活是基于离心的方法（高达 5000g）在血液中心制备 PC 时从全血样本中分离血小板的一个不想要的副作用。在这里，我们评估了基于声的分离设备从全血袋中分离人血小板的效果。通过声过滤分离的血小板的纯度、激活状态和功能与使用已知产生有限血小板激活的参考方案（包括两次连续的软旋转离心（120g 和 1200g））分离的血小板进行了比较。使用自动化血液学分析仪确定血小板浓度和纯度。通过流式细胞术测量表面表达的 P-选择素和激活的整合素 αIIbβ3，评估血小板激活状态和对胶原和凝血酶的反应性。通过氯化铁诱导的血栓形成模型，在活体显微镜下研究了将分离的血小板输注到 NOD/SCID 小鼠时整合到血栓中的能力。通过声过滤血液分离，从血小板部分中消除了超过 80%的红细胞和白细胞，其平均纯度为 92.8±12.8%。声过滤分离的血小板的激活状态和对胶原和凝血酶的反应性与软旋转离心分离的血小板相似。最后，声过滤分离的血小板在氯化铁诱导的血管损伤后被束缚、粘附在血管壁上并整合到生长中的血栓中。总之，我们的结果表明，声过滤是一种用于分离血小板激活最小化和血小板功能保存的人类血小板的合适方法。
