1、血小板低下症的区分及血小板输注合理评估,张志升台湾台大医院输血医学科2016/6/26,Content,血小板低下症的原因新生儿血小板低下症药物诱导血小板低下症血栓性血小板低下症特发性血小板减少紫癜肝素诱导血小板低下症,血小板低下症的原因,骨髓制造血小板的功能减少血小板制造可因放射线、化学治疗或病毒感染等原因受抑制，而骨髓本身疾病，如再生不良性贫血、白血病，骨髓分化不良等皆可引起制造减少。血小板在周边血液被破坏增加常见的有经由产生自体免疫抗体来破坏，如全身性红斑性狼疮、特发性血小板减少性紫斑症、某些药物等，而病毒感染也常引起血小板低下，如登革热病毒、人类免疫缺乏病毒等。此外血小板低下也常见于脾
2、脏肿大病人，如肝硬化病人因门脉高压引起脾脏肿大，造成血小板破坏增加。,Evaluation of a child with thrombocytopenia,Platelet count3 mouths,CBC, blood smear evaluation,Anemia + thrombocytopenia,pancytopenia,Platelet clumps present,pseudothrombocytopenia,Ill appearing?,No,Congenital anomalies?,Yes,No,PMN hypersegmentation RBC macroovaloc
3、ytosis?B12 or RBC folate,B12 or folate deficiency,Medications Immunizations IrradiationToxins?,No,Yes,Macrothrombocytes,Drug-induces Live immunizationIrradiation Toxins,Yes,No,Other morphologic platelet changes,No other platelet change,Bone marrow,Cyanotic heart diseaseFanconi anemiaDyskeratosis con
4、genitaTrisomy 13 or 18Syndromes: Kasabach-Merritt TAR Alport variants,Syndromes:May-hegglinHermansky-PudiakGray platelet,ITPHereditary thrombocytopeniaBernard-Soulier,NI megakaryocytes,megakaryocytes,LeukemiaAplastic anemiaDrug-induced Amegakaryocytic thrombocytopeniaMyelodysplasia,ITP is a diagnosi
5、s of exclusionResponse to therapy, if needed (corticosteroid, IVIG, anti-D antibody), confirms the diagnosis,Yes,PTT, PT, TT,prolonged,DICR/O sepsis,See Consumptional coagulopathy,Normal,SpleenSigns of portal hypertensionplatelet 50,000+/-pancytopenia,MaleEczemaRecurrent infectionSmall platelets,Lym
6、phadenopathyHepatosplenomegalySuperior vena cava syndromeAbdominal mass,Chronically ill appearing,Acute, fibrile illness,WBC enzyme assays UltrasonographyThick smear,Biopsy of lymph node, mass or bone marrow consider tumor lysis and superior vena cava syndromes,HIV assayANAU/ARenal function,Blood cu
7、lture ? antibiotics,MalariaGaucher diseasePortal hypertensionHepatic schistosomiasisCavernoustransformation of the portal vein,Wiskott-Aldrich syndrome,Lymphoma: Hodgkin Non-Hodkin NeuroblastomaleukemiaMyelodysplasia,HIV Autoimmune or connective tissue diseaseHUS/TTP + other microangiopathies Prosth
8、etic cardiac valve,R/O ADMAT-13DATAuto/allo anti-platelet antibody,SepsisVaricellaEBVCMVDenque hemorrhagic feverHIVHUSHantavirusParvovirusOther viruses TTP,Auto/all anti-platelet antibodies study,Heparin-induced thrombocytopenia,Check PF4,Thrombocytopenia in the ill neonate,Any etiology of thrombocy
9、topenia that occurs in the well child,History, examination, CBC, blood smear evaluation,See Thrombocytopenia in the well neonate,Platelets 100,000149,000/uL,Platelets 100,000/uL,If platelets 150,000/uL no further evaluation,100,000149,000 continue to fellow,PTT, PT, TT,High Hb,Severe jaundice and lo
10、w Hb,Prolonged PTT, PT and/or TT +/- microangiopathic hemolytic anemia:Consider D-dimer of FSP, and/or fibrinogen +/- factors II, V and VIII,PolycythemiaCyanotic congenital heart disease,Erythroblastosis fetalisExchange transfusion pphototherapy,DIC,EtologiesAcute infectionAsphyxiaRDSMeconium aspira
11、tionObstetrical complicationsShockThrombosisSevere hemolytic disease of the newbornSevere hepatic disease,TP usually mild enough not to require transfusion except in DIC due to erythroblastosis fetalis,Treat underlying disease Maintain platelets 50,000 with transfusionsMaintain fibrinogen 1.0g/L and
12、 PT WNL with FFP +/- cyrorecipitate,Normal PTT, PT, TT,RDSPulmonary hypertansionMeconium aspirationMechanical ventilation,Infection ViralBacterial Fungal,Perinatal asphyxia,No other specific etiology identified,Unknown etiology,Ongoing re-evaluation if platelets 20,000 in stable full term neonates,
13、50,000 with hemorrhage, surgery, or more extremely preterm infantsObserve for DIC,免疫性血小板低下,Neonatal alloimmune thrombocytopenia (NAIT)Platelet transfusion refractoriness (PTR)Post-transfusion thrombocytopenic purpura (PTP)Passive alloimmune thrombocytopenia (PAT)Transplantation-associated alloimmune
14、 thrombocytopenia (TAATP),Alloantigens implicated in allo immune thrombocytopenia,Dr. N.H. Tsuno presented in 24th regional congress of ISBT,血小板相关抗体疾病或异常,新生儿免疫性血小板减少症NAITP输血后紫斑症PTP血小板输血无效症PTR免疫性血小板低下紫瘢症ITP血栓性血小板低下紫癜TTP肝素刺激血小板低下症HIT药物抗体血小板低下症DIT 严重输血相关呼吸窘迫症候群TRALI,即输血小板两次以上无法达到预期血小板的增加数称为 “ 血小板输注无疗效
15、” 成因: 异体免疫, 自体抗体, ABO血型不符, 病人因素 ( 如急性出血或组织移植排斥.等 ), 药物治疗 ( 如抗生素vancomycin.)所引起血小板输血后的品质评估: 1小时后其 “ 校正血小板增加数 ” 7000, 此方法亦为侦测有无 “ 异体免疫 ” 的间接方式CCI : ( 输血后血小板数-输血前血小板数) X BSA/ 输注血小板量。,血小板输注无效症,与血小板抗体有关之疾病：,Neonatal alloimmune thrombocytopenia 白种人中此病之报告较多，母体之抗血小板抗体进入胎儿内，造成新生儿之血小板异常低下，此病可以生于第一胎。西方人报告中以HPA
16、-1a 抗体为主(又名anti-plA1)，多发生于HPA-1a阴性且HLA-DR3*0101之妇人。日本则曾报告过HPA-4b抗体引起之新生儿血小板减少症 Post-transfusion purpura (PTP) 输血后紫斑症，病人输血后发生血小 板异常降低，引起反应的血品包括血小板，红血球等。部份输血病人体内可以验出血小板抗体，文献报告中最多的也是HPA-1a抗体(anti-plA1 ),Content,血小板低下症的原因新生儿血小板低下症药物诱导血小板低下症血栓性血小板低下症特发性血小板减少紫癜肝素诱导血小板低下症,Neonatal Alloimmune Thrombocytopen
17、ia (NAIT),NAIT due to anti-HLA antibodiesCase of NAIT suspectedly due to anti-HLA are reported, but the association needs to be confirmed.Class I HLA Abs are found in about one third of multiparous women (1531%), and anti-HPA Abs less frequency; however, platelet destruction is usually caused by the
18、 anti-HPA AbsProtective immune mechanism of the placenta : anti-HLA antibodies adsorbed by the stromal cells of placenta expressing paternal antigens; routinely, the infants are born with normal platelet counts.,Dr. N.H. Tsuno presented in 24th regional congress of ISBT,Specificity of HPA Ab in NAIT
19、(Japan),Dr. N.H. Tsuno presented in 24th regional congress of ISBT,The risk of ICH was the highest with anti-HPA-3,血小板抗体的特异性70 NAITP案例,Dr. G.G. Wu presented in 24th regional congress of ISBT,2013 ISBT IPIWP-AR,The risk of ICH was the highest with anti-HPA-3Anti-HPA-4a/4b is the highest rate in NAIT
20、in Japan ( anti-HPA-4a 7%, anti-HPA-4b 52%) HPA-4 incompatible may cause rejection rection reactions in solid organ transplantation.(Vox Sanguinis, 2010 july, Supplement1,Vol 99)More than 0.5% of CD36 type I deficient individuals are at risk to be immunized through blood transfusion or pregnancy in
21、China.,The frequency of CD36 negative,新生儿免疫血小板低下症(NAIT),NAIT due to anti-HLA antibodiesCase of NAIT suspectedly due to anti-HLA are reported, but the association needs to be comfirmedClass I HLA Abs are found in about one third of multiparous women (1531%), and anti-HPA Abs less frequently; however,
22、 platelet destruction is usually caused by the anti-HPA Abs.Protective immune mechanism of the placenta: anti-HLA antibodies adsorbed by the stromal cells of placenta expressing paternal antigens; routinely, the infants are born with normal platelet counts.,新生儿血小板低下的评估(NATP),新生儿免疫血小板低下症的评估诊断,母体血清验血小
23、板抗体PIFTELISA法母体血清和患儿血小板交叉SPRCA, FCM母体血清和父亲血小板交叉SPRCA, 流式细胞仪,This assay can detect HLA, ABO, and platelet specific IgG antibodies. But weak reactions give immediate results.,Solid- phase red cell adherence assay (SPRCA),输血后紫斑症,Post-transfusion thrombcytopenia purpuraDeveloped after 1-2 weeks after tr
24、ansfusionsCaused by Anti-HPA -1a,Content,血小板低下症的原因新生儿血小板低下症药物诱导血小板低下症血栓性血小板低下症特发性血小板减少紫癜肝素诱导血小板低下症,药物诱导血小板低下症抗体,大部份药物诱导血小板低下的症状轻微，但偶而有危及生命的出血状况产生。病理原因较药物诱导红细胞溶血性贫血复杂，至少有六种以上的机转主要临床意义在于排除血小板低下原因,J Thromb Haemost. 2009 Jun; 7(6): 911918.,DIT药物诱导血小板低下抗体鉴定,血小板交叉配血相容，1Hr CCI7500, 24 Hr CCI5000 有效输血小板1Hr
25、CCI7500, 24 Hr CCI5000 infection, fever, non-immune1Hr CCI7500, 24hrs CCI5000,配血相容, 1 hr CCI7500, 24hrs CCI5000,配血相容, 1 hr CCI7500,配血不相容, 1 hr CCI7500,执行血小板抗体筛检HPA or HLA class I Ab, or CD36,发烧等非免疫因素,成功输血,药物诱导型血小板低下,跟临床取得药物及查询药物作用浓度,配制药物浓度, 执行药物依赖型检测SPRCA,更改药物,药物诱导型血小板低下抗体鉴定案例,男性68岁大肠癌患者，使用oxaliplas
26、tin合并5-FU化疗使用，每周一次疗程，约使用三星期疗程后发觉血小板从原来17万/dL, 不明原因降为5.8万/dL, 无出血现象，也无其他败血症及DIC等情形。其他检查：凝血功能PT/aPTT正常，血小板交叉配血相容，血小板抗体筛检Pakplus ELISA酶标法无血小板抗体。输血后一小时CCI 6000.,Negative,Weak positive,Positive,疑似药物诱导溶血性贫血与药物诱导血小板低下出血案例,一居住在台湾东部花莲原住民族男性45岁酒精性肝炎住院患者。之前的病史及输血史：A型Rh阳性，年轻时(30岁左右因工作受伤)手术曾输过RBC 6U及1U机采血小板。三四年前
27、曾因急性肺炎住院接受治疗，长期使用抗生素(泰巴坦)治疗此次住院治疗疲倦黄疸，有伤口长期无法愈合不良合并有肾结石发烧出血等症状实验室检查A/pos 抗筛阴性，Hb 6.5, platelet count 50k, aPTT/PT prolong INR 1.7 blood culture no grow 建议抗生素及输血治疗,疑似DIHA合并DIT输血治疗案例,临床医师建议输给RBC 4U, 机采血小板1U, FFP 6U及泰巴坦治疗, 抗筛阴性输血科给予随机A型RBC相叉相容RBC 4U 及A型机采血小板1U, FFP6U 输注。输血后第一天Hb 7.2 直抗转阳, Plt 45K 溶血三症明
28、显，LDH，疑输血后溶血症，输血科启动输血反应调查SOP。输血科输血反应调查：输血后样本抗筛阴性，直抗转阳，放散液抗筛阴性和原献血者样本交叉相容，重复输血后样本对照输血前样本交叉配血，输血后样本的主侧配血AHG有1+凝集，凝聚胺配血相合，输血前样本配血均相容。临床医师建议再次输血RBC 4U, Plt 1机采血小板.输血科再次配血相容RBC 4U及1U 机采血小板输注。输血后第二天溶血评估加剧，疑似引发DIC ( D-dimer, FDP上升) Hb5.0, plt 8k, 无大量出血现象.,疑似DIHA合并DIT输血治疗案例-cont.,输血科再次启动输血反应调查作业，同时间病患出现EV b
29、leeding 输血科紧急供应交叉配血相容RBC 及机采血小板, FFP, Cryo。最后输血不及, 病患死亡事后调查: 血小板抗体筛检阴性( Genprobe Pakplus ELISA), auto-platelet Ab.阳性( FIPA, Flowcytometry流式细胞仪)DAT : polyAHG 3+, IgG AHG 3+, C3d negative, 放散液抗筛阴性重测抗筛阴性, 红细胞主交叉: 相容.,DITP的治疗,立即停药；血小板输注；大剂量IVIG；短期使用糖皮质激素；避免再次使用该药物,Content,血小板低下症的原因新生儿血小板低下症药物诱导血小板低下症血栓性
30、血小板低下症特发性血小板减少紫癜肝素诱导血小板低下症,栓塞性血小板低下紫斑症TTP,形成的原因仍不清楚，多数学者认为是一种病毒传染后毒素所造成的反应。但临床上可发觉正常金属蛋白酵素（metalloprotease,ADAMTS-13）可以分解超大von Willebrands体。它具有类似thrombospondin-1单元（thrombospondin-1like domains），并藉此与内皮细胞上的thrombospondin接受体结合，并由此固定于内皮细胞上。固定于内皮细胞上的ADAMTS 13，使可以分解旁边的超大Von Willebrands氏因子聚合体。栓塞性血小板低下紫斑症患者
31、，其金属蛋白酵素（metalloproteaseADAMTS 13）于此时的活性若严重通常趋近于零，无法分解旁边的超大Von Willebrands氏因子聚合体，所导致的微血管内血小板凝集，因而表现出所谓的pentad：包括微血管病变溶血性贫血、血小板低下、发烧、神经学症状、以及肾功能不全等五种特征。,The ADAMTS-13 assay is based on fluorescence resonance energy transfer (FRET) technology. A synthetic fragment of the von Willebrand Factor protein
32、is used as the Substrate. Cleavage of this peptide between two modified residues releases the fluorescence quenching capabilities.This assay is based on quantifying the cleavage of a small fragment of von Willebrand Factor by the ADAMTS-13 protease. The cleavage of this synthetic substrate is detect
33、ed by reading the fluorescence that results when the substrate is cleaved.,测定ADAMTS-13的原理,TECHNOZYM ADAMTS-13 ACTIVITY,PE对TTP的疗效,一般血浆交换术PE对TTP的临床反应可谓是十分之迅速且明确。Bukowski报告2-3天，Petitt报告约36小时。血小板约25 PE可见成效，但血色素较略显缓慢。LDH恢复则较慢。对不同病因的TTP之临床反应速度快慢差异颇大，有约连续5天疗程即完成疗效，有近一个月或以上连续QD PE才完成疗效。以大量血浆新鲜血浆，新鲜冷冻血浆，冷冻血浆
34、补充效果较佳，开始PE疗程QD实施，超过一个blood volume的血浆置换术急性期治疗常使用120%，以platelet count评估成效。,PE & PI 对TTP的疗效,Plasma infusion (PI)bas been used as an alternative to PE.Studies observed no difference in response or survival between PE and PI.The risks of fluid overload with PI and the potential for PE to remove ADAMTS13
35、 have resulted in PE being preferred to PI,TTP之血浆疗法,自1977年血浆疗法的临床应用已显着地改善血栓性血小板减少性紫斑症(TTP)之愈后。使用血液成份分离机及血浆分离术或者是血浆输注法plasma infusion可治愈大部分的TTP病人。唯此两种血浆疗法之相互优劣比较及引发TTP之致病假说，仍尚无定论。1997年新光医院温武庆/叶建宏等发表一例TTP，交互使用双重过滤血浆分离术DFPP配合血浆输注疗法，及血浆交换术的案例报告：初期以连续16次DFPP配合每天之血浆输注疗法，病人之临床征状却依然持续恶化，只得改用传统血浆交换术。结果在二天之内血
36、小板数目及神智状态明显进步。比较此二种血浆疗法之个别差异，发现血浆交换术中每次使用之血浆量大约是血浆输注法之三倍，因此，大量的血浆输注应是治疗成功的主因。台湾医学 Formosan J Med 1997;6:710-5,Cryo-poor plasma(cryosupernatant),laboratory indices of complete and stable response (platelet count, serum lactate dehydrogenase level) did not normalize in concert with clinical improvemen
37、t.,PE of TTP in NTUH,2006.01.2008计十五名疑似TTP如下一张slide。其中一名E先生，症状十分疑似，但ADAMTS-13测出达80%normal，执行四次PE换血浆64U后停止PE治疗，使用传统platelet transfusion可回复到180k。其他十二名, 男3名平均52.5岁，女9名平均32.1岁H女士只执行一次PE，临床症状十分吻合，原高度怀疑为TTP，但ADAMTS-13测出达78%最后确认是Trousseaus syndrome 。,20052008 TTP案例报告,ADAMTS-13对TTP的鉴别,H女士：Poor prognosis was
38、 informed. Pupil dilate, unconsciousness and no gag reflex were found. Held PLT transfusion due to suspect TTP. FFP was kept as hematologist suggestion. However, vaginal bleeding and oral bleeding persisted. The ADAMT13 was about 80%. TTP was not likely. Elevated CA 125 was found.The abdomen CT show
39、ed ovarian cancer and endometrial hyperplasia. Trousseaus syndrome was suspected.,ADAMTS-13对TTP的鉴别(二),某君从新店慈济转入本院急诊，疑似TTP的案例。临床症状及实验室检查疑似TTP：LDH，Bilirubin，Hb，Platelet,Coombstest: negative, unconsciousness ,唯PT, aPTT prolong,留检体评估ADAMTS-13。病人随即CPR，ADAMTS-13 assay: 5%,suspected terminal TTP.,血栓性血小板低下的
40、区分(TTP),Content,血小板低下症的原因新生儿血小板低下症药物诱导血小板低下症血栓性血小板低下症特发性血小板减少紫癜肝素诱导血小板低下症,ITP特发性血小板减少症,ITP的治疗主要是在降低临床重大出血的风险。严重出血的ITP患者，输血仍是最直接有效的，但缺乏评估输血1小时后的有效的血小板数的循证IVIG 1g/kg (5000/mL, 每日追踪)Glucocorticoids (1g IV, 每日3dose),ITP的实验室诊断,取auto platelet 测auto antibodyELISA PakAuto kit, flowcytometry, SPRCA最近无施打IVIG,
41、 无输过血小板，采SPRCA: ,50000 plat count, 抽40cc EDTA，制成PRP,This assay can detect HLA, ABO, and platelet specific IgG antibodies. But weak reactions give immediate results.,Solid- phase red cell adherence assay (SPRCA),Content,血小板低下症的原因新生儿血小板低下症药物诱导血小板低下症血栓性血小板低下症特发性血小板减少紫癜肝素诱导血小板低下症,肝素介导的血小板减少（HIT）,机制： 肝素与
42、血小板因子4(PF4)结合形成抗原结构，抗体通过Fab段识别PF4/肝素结合于血小板，Fc段连接单核细胞，迅速、猛烈地引起血小板激活以及促凝微粒的释放。类似机制的药物：鱼精蛋白 鱼精蛋白-肝素-IgG-血小板FcRIIa,EDTA依赖假性血小板低下Vs.HIT,HIT的抗体侦测：血清素释放试验serotonin release assay SRA(高特异性,低敏感性), 抗体-肝素-PF4免疫复合体试测酶标法(高敏感性低特异性性),血小板输血参考指标IPF,IPF是Immature platelet fraction(未成熟血小板比例), 可作为评估骨髓造血小板功能恢复的参考指标化疗或干细胞移
43、植后, IPF开始上升代表骨髓造血小板功能逐渐恢复，约一周内血小板会显着上升。降低不必要的血小板输血,IPF与造血小板功能,IPF可早期评估骨髓造血小板功能,评估停上血小板输血时机降低输血成本降低输血造成感染的风险,美国红十字会与英国输血组织移植联盟血小板配型流程,Crossmatch with 8 random donors,All donors negative: Use random donors for PLT Tx,8 Crossmatch positive,Positive and negative donors,Select negative donor for PLT TX,H
44、LA/HPA antibody test,Anti-HPA +Anti-HLA -,Anti-HPA -Anti-HLA +,Anti-HPA +Anti-HLA +,HPA typing of recipient,HLA-identical or compatible PLT,Crossmatch with HPA compatible donor,HPA typing of recipient,Crossmatch with HLA/HPA compatible donor,二次以上输随机血小板无效,HLA class I 筛查,positive,Negative或持续对HLA相容血小板输注无效,病患的HLA-I分型,提供HLA相容配型血小板,持续对HLA及HPA配血相容血小板输注无效,考虑非免疫问题或药物诱导型抗体或HPA专一性筛检,阳性,阴性,执行HPA抗体筛检,提供HPA相容配型血小板,区分血小板相关疾病,区分各种血小板低下ITP, PTR, TTP, HIT, DIT,
