一次离心法制备大、小鼠富血小板血浆实验条件的优化
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摘要
目的探讨一次离心法制备大、小鼠富血小板血浆(PRP)的最佳离心条件。方法分别采用股动脉插管和心脏穿刺的方法获取大、小鼠动脉血,14%CPDA-1抗凝,滤除白细胞后分装入无菌EP管中,不同条件下(离心力:300×g～600×g,离心时间:4～12min)离心制备大、小鼠PRP,利用血液分析仪分别对抗凝全血、滤除白细胞后的血样及PRP进行细胞计数,比较不同方法间的血小板回收率。结果大、小鼠抗凝全血滤除白细胞后分别以400×g和300×g的离心力离心8min时,血小板回收率最高。结论选择合适的离心力和离心时间,达到白膜形成前的临界状态,是一次离心法制备PRP的关键。
Objective To investigate the optimal centrifugation conditions for preparation of rat and mouse platelet rich plasma(PRP)by single centrifugation.Methods Arterial blood of rats and mice by femoral artery cannulation and cardiac puncture were obtained respectively,anticoagulation with 14% CPDA-1,while white blood cells in the blood were filtered out.Then the blood was divided into sterile EP tubes,while PRP was prepared by centrifugation in different conditions(the centrifugal force was 300×g-600×g,and the centrifugal time was 4-12 min).The number of blood cells of the anticoagulant whole blood,the leukocyte-depleted blood sample and PRP were counted by hematology analyzer,and platelet recovery rates were compared between different methods.Results The platelet recovery rate was highest when the blood samples of rats and mice were centrifuged at 400×g and 300×g for 8 min respectively.Conclusion It is a key to prepare PRP by single centrifugation that selecting the appropriate centrifugal force and time and reaching a critical state before the formation of the buffy coat.
Objective To investigate the optimal centrifugation conditions for preparation of rat and mouse platelet rich plasma(PRP)by single centrifugation.Methods Arterial blood of rats and mice by femoral artery cannulation and cardiac puncture were obtained respectively,anticoagulation with 14% CPDA-1,while white blood cells in the blood were filtered out.Then the blood was divided into sterile EP tubes,while PRP was prepared by centrifugation in different conditions(the centrifugal force was 300×g-600×g,and the centrifugal time was 4-12 min).The number of blood cells of the anticoagulant whole blood,the leukocyte-depleted blood sample and PRP were counted by hematology analyzer,and platelet recovery rates were compared between different methods.Results The platelet recovery rate was highest when the blood samples of rats and mice were centrifuged at 400×g and 300×g for 8 min respectively.Conclusion It is a key to prepare PRP by single centrifugation that selecting the appropriate centrifugal force and time and reaching a critical state before the formation of the buffy coat.
引文
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[18]XIA S,CHEN G,WANG B,et al.Addition of sodium pyruvate to stored red blood cells attenuates liver injury in a murine transfusion model[J].Mediators Inflamm,2016,2016:3549207.
[19]TAO Z Y,XIA H,CAO J,et al.Development and evaluation of a prototype non-woven fabric filter for purification of malaria-infected blood[J].Malar J,2011,10:251.
[2] PIETRAMAGGIORI G,SCHERER S S,MATHEWS J C,et al.Quiescent platelets stimulate angiogenesis and diabetic wound repair[J].J Surg Res,2010,160(1):169-177.
[3] MARIA-ANGELIKI G,ALEXANDROS-EFSTRATIOS K,DIMITRIS R,et al.Platelet-rich plasma as a potential treatment for noncicatricial alopecias[J].Int J Tricol,2015,7(2):54-63.
[4] MOGHADAM A,KHOZANI T T,MAFI A,et al.Effects of platelet-rich plasma on kidney regeneration in gentamicin-induced nephrotoxicity[J].J Korean Med Sci,2017,32(1):13-21.
[5] MARTN-SOLO,RODJ,GARCA-APARICIO L,et al.Effects of platelet-rich plasma(PRP)on a model of renal ischemia-reperfusion in rats[J].PLoS One,2016,11(8):e0160703.
[6] SEKERCI C A,TANIDIR Y,SENER T E,et al.Effects of platelet-rich plasma against experimental ischemia/reperfusion injury in rat testis[J].J Pediatr Urol,2017,13(3):317.
[7] JANSEN A J,JOSEFSSON E C,RUMJANTSEVA V,et al.Desialylation accelerates platelet clearance after refrigeration and initiates GPIbαmetalloproteinase-mediated cleavage in mice[J].Blood,2012,119(5):1263-1273.
[8] ZHAO J,SUN Z,YOU G,et al.Transfusion of cryopreserved platelets exacerbates inflammatory liver and lung injury in a mice model of hemorrhage[J].J Trauma Acute Care Surg,2018,85(2):327-333.
[9] LORENZO-ZUIGA V,BOIX J,MORENO DE VEGA V,et al.Efficacy of platelet-rich plasma as a shielding technique after endoscopic mucosal resection in rat and porcine models[J].Endosc Int Open,2016,4(8):E859-864.
[10]K L,GNAY H,ERBAS O,et al.Effects of platelet-rich plasma on nerve regeneration in a rat model[J].Acta Orthop Traumatol Turc,2014,48(4):449-454.
[11]MAMMOTOT,JIANGA,JIANGE,et al.Platelet-rich plasma extract prevents pulmonary edema through angiopoietin-Tie2signaling[J].Am J Respir Cell Mol Biol,2015,52(1):56-64.
[12]STEFANINI L,RODEN R C,BERGMEIER W.CalDAGGEFI is at the nexus of calcium-dependent platelet activation[J].Blood,2009,114(12):2506-2514.
[13]HOSHI R,MURATA S,MATSUO R,et al.Freeze-dried platelets promote hepatocyte proliferation in mice[J].Cryobiology,2007,55(3):255-260.
[14]DOLE V S,BERGMEIER W,MITCHELL H A,et al.Activated platelets induce Weibel-Palade-body secretion and leukocyte rolling in vivo:role of P-selectin[J].Blood,2005,106(7):2334-2339.
[15]ZHANG J,YUAN T,ZHENG N,et al.The combined use of kartogenin and platelet-rich plasma promotes fibrocartilage formation in the wounded rat Achilles tendon entheses[J].Bone Joint Res,2017,6(4):231-244.
[16]DHURAT R,SUKESH M.Principles and methods of preparation of platelet-rich plasma:a review and author′s perspective[J].J Cutan Aesthet Surg,2014,7(4):189-197.
[17]ARGELLES D,CARMONA JU,PASTOR J,et al.Evaluation of single and double centrifugation tube methods for concentrating equine platelets[J].Res Vet Sci,2006,81(2):237-245.
[18]XIA S,CHEN G,WANG B,et al.Addition of sodium pyruvate to stored red blood cells attenuates liver injury in a murine transfusion model[J].Mediators Inflamm,2016,2016:3549207.
[19]TAO Z Y,XIA H,CAO J,et al.Development and evaluation of a prototype non-woven fabric filter for purification of malaria-infected blood[J].Malar J,2011,10:251.