目的:探讨n JAK2 V617F基因突变呈阴性原发性血小板增多症(ET)患儿的临床、实验室检查及基因突变特征。n 方法:选择2017年2月至2019年2月于首都医科大学附属北京儿童医院血液肿瘤中心接受诊治的10例n JAK2 V617F基因突变呈阴性ET患儿作为研究对象，纳入ET组(n n＝10)。患儿中位年龄为6.8岁；男性患儿为5例，女性为5例。选择同期于本院接受诊治的4例继发性血小板增多症(ST)患儿作为对照，纳入ST组(n n＝4)。患儿中位年龄为1.8岁，均为女性。采用病例对照的研究方法，分析2组患儿的血常规、凝血功能检查、骨髓涂片、骨髓活组织检查及相关基因突变检查结果。2组患儿年龄、血常规和凝血功能检测结果、巨核细胞总数及其比例的比较，采用Mann-Whitney n U检验。性别、临床表现构成比比较，采用Fisher确切概率法。本研究遵循的程序符合2013年修订的《世界医学协会赫尔辛基宣言》要求。所有患儿临床资料收集均已获得患儿监护人知情同意。n 结果:① ET组10例患儿中，伴神经系统症状者为6例，包括头痛者4例、头晕2例；其中1例同时伴肝大；其余4例无明显临床症状。4例ST组患儿中，伴发热症状或者前驱感染者为2例，其余2例无明显临床症状。② 2组患儿外周血血小板计数、血小板压积(PCT)均较正常参考值范围增高，ET组患儿中位血小板计数为1 451×10n 9/L[(1 193~1 831)×10n 9/L]，高于ST组患儿的966×10n 9/L[(677~989)×10n 9/L]，并且差异有统计学意义(n Z＝-2.404，n P＝0.014)；中位PCT为1.28%(1.12%~1.63%)，亦显著高于ST组患儿的0.85%(0.62%~1.05%)，并且差异有统计学意义(n Z＝-2.256，n P＝0.024)。2组患儿的血小板分布宽度(PDW)均低于正常参考值范围。凝血功能相关指标中，ET组患儿中分别有4、4、4例出现凝血酶原时间(PT)、部分凝血活酶时间(APTT)延长及纤维蛋白原(FIB)值低于正常参考值范围，而ST组患儿中分别为1、2、2例。2组患儿其余检查指标均正常。③ 2组患儿骨髓涂片结果均表现为中度增生活跃，粒、红系细胞比例正常。此外，4例ST组患儿的骨髓涂片中亦可见中性粒细胞核左移和感染相关细胞。2组患儿的骨髓活组织检查结果均表现为中度增生活跃，粒、红系细胞比例正常且巨核细胞多见，以分叶核巨核细胞为主。巨核细胞免疫组织化学染色结果示，ET组患儿骨髓中位正常巨核细胞比例显著低于ST组患儿[49.5%(44.3%~61.0%)比71.1%(61.3%~85.0%)，n Z＝-2.404，n P＝0.014]。④基因测序结果显示，ET组10例患儿均不伴n MPL及n JAK2基因突变，其中有2例检测到n CALR基因突变，并且这2种n CALR基因突变均为明确与ET相关的基因突变位点。此外，筛选出4种可能与儿童ET相关的基因突变，包括n KMT2A、n ASXL1、n CSF3R及n NF1基因突变。n 结论:JAK2 V617F基因突变呈阴性ET患儿具有独特的临床和实验室特征，二代基因测序法发现可能与ET发生、发展相关的n KMT2A、n ASXL1、n CSF3R及n NF1基因突变，可为n JAK2 V617F基因突变呈阴性ET的临床诊断提供参考。但是本研究纳入患儿例数较少，并且缺乏ST男性患儿作为对照，存在一定局限性。ET患儿的临床特点、相关实验室检查结果，及其与ST患儿的鉴别要点，有待大样本临床研究进一步证实。n “,”Objective:To investigate clinical, laboratory examination and gene mutation characteristics of n JAK2 V617F gene mutation negative essential thrombocytosis (ET) children.n Methods:A total of 10 cases of n JAK2 V617F gene mutation negative ET children who were treated in Hematological Oncology Center, Beijing Children′s Hospital, Capital Medical University from February 2017 to February 2019 were included as research subject and enrolled into ET group (n n=10). Median age of children was 6.8 years. There were 5 males and 5 females. Four children with secondary thrombocytopenia (ST) who received treatment in our hospital during the same period were included as control and enrolled into ST group (n n=4). Median age of children was 1.8 years. All the children were female. Blood routine, coagulation function test, bone marrow smear, bone marrow biopsy and related gene mutation test results of children in two groups were analyzed through case-control study mothod. Mann-Whitney n U test was used to compare the age, results of blood routine examination, values of coagulation function, total numbers and proportions of megakaryocytes in two groups. Fisher probabilities was used to compare composition ratios of gender and clinical manifestations. This study met requirements of n World Medical Association Declaration of Helsinki revised in 2013. And all clinical data of the children have been collected with informed consent of children′s guardians.n Results:① Six of 10 children in ET group had neurological symptoms, including headache in 4 cases and dizziness in 2 cases. One patient had concurrent hepatomegaly. Remaining 4 cases had no obvious symptoms. Among 4 children in ST group, two cases were accompanied by fever or prodromal infection, and the other 2 cases had no obvious symptoms. ② Peripheral blood platelet count and plateletocrit (PCT) of children in two groups were significantly higher than normal reference range.Median platelet count of children in ET group was 1 451×10n 9/L[(1 193-1 831)×10n 9/L], which was higher than that of children in ST group, which was 966×10n 9/L[(677-989)×10n 9/L], and difference was statistically significant (n Z=-2.404, n P=0.014). And median PCT of children in ET group was 1.28%(1.12%-1.63%), which was also higher than 0.85%(0.62%-1.05%) of ST group, and difference was statistically significant (n Z=-2.256, n P=0.024). Platelet distribution width (PDW) of children in two groups were lower than the normal reference range. Among the coagulation function related indicators, prothrombin time (PT), activated partial thromboplastin time (APTT) prolongation and fibrinogen (FIB) reduction were observed in 4, 4, 4 children in ET group, while in 1, 2, 2 children in ST group, respectively. Other examination indicators of children in two groups were normal. ③ Bone marrow smear results of children in two groups showed moderate hyperplasia, and ratio of granulosa to red system was normal. In addition, left shift of neutrophil nuclei and infection-related cells were also observed in bone marrow smears of 4 children in ST group. Bone marrow biopsy results of children in 2 groups showed moderate hyperplasia and active hyperplasia, and grain red ratio was normal. Megakaryocytes were dominated by lobulated megakaryocytes. Megakaryocyte immunohistochemical staining showed that median proportion of normal megakaryocytes in bone marrow of children in ET group was significantly lower than that of children in ST group [49.5% (44.3%-61.0%) n vs 71.1% (61.3%-85.0%), n Z=-2.404, n P=0.014]. ④ Gene sequencing showed that none of 10 children in ET group had n MPL or n JAK2 gene mutations, and 2 of them had n CALR gene mutations, which were clearly related to ET. In addition, four gene mutations that may be associated with ET in children were screened, including n KMT2A, n ASXL1, n CSF3R and n NF1 gene mutation.n Conclusions:JAK2 V617F gene mutation negative ET children have unique clinical and laboratory examination characteristics. Next-generation sequencing found mutations in n KMT2A, n ASXL1, n CSF3R and n NF1 genes that may be related to occurrence and development of ET, which can provide reference for clinical diagnosis of n JAK2 V617F gene mutation negative ET. However, the number of children included in this study was small, and there was a lack of male children with ST as a control, which had certain limitations. Clinical characteristics of n JAK2 V617F gene mutation negative ET children, results of relevant laboratory tests, and key points of differentiation from ST children need to be further confirmed by large-sample clinical studies.n