真性红细胞增多症常用诊断标准比较

目的:描述3种真性红细胞增多症(PV)诊断标准:国内诊断标准、WHO2008年诊断标准和BCSH诊断标准,并比较3种诊断标准的敏感性及特异性。方法:统计50例近期在中国医科大学附属盛京医院就诊以红细胞增多为主要临床表现的病例,自就诊以来相关信息,根据病史及临床疗效将患者分组,将此结果与3种诊断标准得出的结果进行对比。结果:在纳入分析范围的45例患者中,根据病史及临床疗效将35例诊断为PV,其余10例为继发性红细胞增多。将国内诊断标准、WHO诊断标准及BCSH诊断标准分别与临床诊断结果对比,得到3种诊断标准的敏感性分别为51.43%、85.71%和91.43%,特异性分别为100%、70%和90%。结论:JAK2V617F基因突变在PV诊断中有重要地位。在JAK2V617F基因突变阴性的情况下,BCSH诊断标准较为精准;而当JAK2V617F基因突变阳性时,单纯依靠BCSH或WHO诊断标准会造成特异性降低,需要综合参考血小板、中性粒细胞计数和血清Epo水平以提高诊断的准确性和特异性。     

bcr-abl阴性骨髓增生性疾病患者JAK2V617F点突变检测的临床意义

目的 探索Janus Kinase 2V617F基因突变(JAK2V617F)在bcr-abl阴性骨髓增生性疾病(MPD)的发生率和临床意义.方法 基因组DNA从患者骨髓或外周血粒细胞中提取.采用等位基因特异性PCR(AS-PCR)、限制性内切酶消化和PCR产物测序的方法检测JAK2V617F突变.共检测患者110例,其中bcr-abl阴性MPD 41例、bcr-abl阳性慢性粒细胞白血病(CML)25例和急性白血病44例.结果 JAK2V617F阳性结果分别为真性红细胞增多症(PV)11例(91.7%)、原发性血小板增多症(ET)8例(53.3%)、特发性骨髓纤维化(IMF)4例(57.1%),而高嗜酸粒细胞增多症(HES)7例、bcr-abl阳性CML 25例和急性白血病44例[包括急性髓细胞白血病(AML)24例、急性淋巴细胞白血病(ALL)18例、急性混合细胞白血病2例1均未检测到JAK2V617F基因突变.在所有JAK2V617F阳性的标本和10份阴性标本中,AS-PCR和限制性内切酶消化方法的测定结果都可经测序进一步证实.结论 90%以上的PV、50%以上的ET和IMF可检测到JAK2V617F基因突变;JAK2V617F基因突变在PV、ET和IMF的诊断和鉴别诊断中有重要意义,可以作为诊断的分子标志,也可能是治疗的新靶点.     

潜匿性真性红细胞增多症与真性红细胞增多症鉴别诊断

目的：对比分析潜匿性真性红细胞增多症（mPV）和真性红细胞增多症（PV）的特点,探讨mPV 早期诊断的方法。方法收集符合诊断标准的男性初诊 PV、mPV 患者各100例,均行促红细胞生成素（EPO）、中性粒细胞碱性磷酸酶（NAP）、 JAK2 V617F 基因检测及骨髓组织活组织检查（BMB）。半年后追踪未经特殊治疗两组患者的血红蛋白（Hb）及 JAK2 V617F 基因突变负荷量变化情况。结果PV 组与 mPV 组 EPO 分别为（3.4±0.7）、（3.2±0.6） U／ml,NAP 积分分别为（276±20）、（278±21）分, BMB 造血容积分别为（78±10）%、（76±9）%,巨核细胞数分别为（53±6）、（51±5）个／张切片,差异均无统计学意义（均 P＞0.05）。 JAK2 V617F 基因突变负荷量分别为（89.2±9.4）%、（78.1±8.6）%,差异有统计学意义（P＜0.05）。 mPV 组中未经特殊治疗的半年后达 PV 水平（Hb≥185 g／L）的37例患者 Hb 平均水平为（194±8）g／L,JAK2 V617F 负荷量为（90.7±9.1）%。结论 PV、mPV 患者的 EPO、NAP 积分及骨髓组织形态学无差异,而 JAK2 V617F 基因突变负荷量有差异。未经特殊治疗的 mPV 患者 Hb 水平半年后多数可达到典型 PV 的诊断水平,JAK2 V617F 基因突变负荷量随之增加。     

BCR-ABL阴性骨髓增殖性肿瘤JAK2 V617F基因突变状态及负荷的临床意义

目的探讨JAK2 V617F基因突变状态及负荷对BCR-ABL阴性骨髓增殖性肿瘤(MPN)的影响。方法回顾性分析2015年9月至2020年1月河北省沧州市中心医院199例MPN患者的临床资料。分析JAK2 V617F突变负荷与MPN患者临床病理特征及预后评分的关系。结果199例BCR-ABL阴性MPN患者中JAK2 V617F突变阳性138例(69.4%);其中,72例真性红细胞增多症(PV)患者中突变阳性64例(88.9%),101例原发性血小板增多症(ET)患者中突变阳性54例(53.5%),25例骨髓纤维化(MF)患者中突变阳性20例(80.0%),1例嗜酸粒细胞增多症(HES)患者突变阳性。JAK2 V617F突变高负荷者占55.1%(76/138)。突变负荷最高的类型为PV,MF次之,ET最低,3组突变负荷分别为(73.9±18.3)%、(59.9±25.2)%、(25.0±16.5)%。JAK2 V617F突变负荷与PV、ET、MF患者的白细胞计数均呈正相关(r值分别为0.626、0.675、0.796,均P<0.01)。JAK2 V617F突变负荷与PV、ET患者的预后评分均呈正相关(r值分别为0.296、0.404,均P<0.05)。结论BCR-ABL阴性MPN患者JAK2 V617F突变负荷与临床病理因素相关,JAK2 V617F突变高负荷患者预后不良。     

真性红细胞增多症27例临床分析

目的 对真性红细胞增多症（Pv）患者的临床特征进行分析,进一步了解其特点,为临床诊治提供帮助.方法 对2001年5月至2012年12月收治并确诊为PV的27例患者的临床资料进行回顾性分析.结果 对27例患者中合并血栓性疾病15例（55.6％）,行JAK2 V617F基因突变检测15例中,12例出现阳性,阳性率为80.0％.结论 JAK2 V617F基因突变在PV患者中的发生率高,对PV早期诊断、治疗及预防血栓事件的发生具有重要的临床意义.     

血红蛋白及血小板增多患者的JAK2 V617F突变检测及临床意义分析

骨髓增殖性肿瘤（myeloproliferative neoplasm,MPN）是以一系或多系分化相对成熟的骨髓细胞克隆性增殖异常为特点的疾病,其分类中的真性红细胞增多症（polycythemia vera,PV）、原发性血小板增多症（essentialthrombocythemia,ET）等疾病中存在JAK2V617F点突变。有研究表明,在检测MPN患者JAK2V617F突变中,PV发生率在90％以上、ET发生率为50％-70％。JAK2V617F基因突变已作为诊断MPN的必要指标之一。本研究目的是对血红蛋白及血小板增多临床怀疑MPN者进行JAK2V617F突变检测,并结合临床资料进行分析,为血红蛋白及血小板增高患者的诊断提供依据。     

骨髓增殖性肿瘤120例JAK2 V617F基因突变的临床分析

目的 探讨JAK2 V617F基因突变在骨髓增殖性肿瘤(MPN)患者中的发生率及临床意义.方法 采用骨髓细胞学和活组织检查方法分析120例患者的骨髓病理状况,监测费城染色体(Ph染色体)和bcr-abl融合基因.从患者骨髓抽提DNA,采用荧光定量PCR技术检测JAK2 V617F基因突变.结果 所有患者均呈现出MPN各自类型的典型特征.Ph染色体和bcr-abl融合基因检测均为阴性.120例MPN患者中JAK2 V617F基因突变的阳性率为66.7％(80/120),其中真性红细胞增多症(PV)为72.7％(16/22),原发性血小板增多症(ET)为66.0％(62/94),4例原发性骨髓纤维化(PMF)患者中2例阳性.JAK2 V617F突变阳性PV患者的外周血白细胞计数(P=0.001)和血小板计数(P=0.010)均高于阴性患者;JAK2 V617F突变阳性ET患者的白细胞计数高于阴性患者(P=0.006);PMF中JAK2V617F突变阳性和阴性患者间各项指标差异均无统计学意义(均P＞0.05).结论 JAK2 V617F基因突变检测有助于bcr-abl阴性MPN的诊断和鉴别诊断,使患者能够在早期被发现和治疗.     

实时定量聚合酶链反应检测慢性骨髓增殖性疾病患者JAK2 V617F基因突变

 目的　观察实时定量聚合酶链反应（PCR）检测慢性骨髓增殖性疾病（CMPD）中JAK2基因V617F点突变情况。方法　采用实时定量PCR法检测56例CMPD患者JAK2 V617F基因突变类型及突变转录水平。其中,真性红细胞增多症（PV）26例、原发性血小板增多症（ET）24例、原发性骨髓纤维化（CIMF） 5例、高嗜酸粒细胞综合征（HES）1例。结果　56例CMPD患者JAK2 V617F基因突变率为51.79 ％（29／56）,PV 65.38 %（17／26）、ET 37.50 %（9／24）、IMF 60.00 %（3／5）;杂合性突变率为41.07 %（23／56）,包括PV 53.85 %（14／26）、ET 29.17 %（7／24）、CIMF 40.0 %（2／5）;纯合性突变率为10.71 %（6／56）, 包括PV 11.54 %（3／26）、ET 8.33 %（2／24）、CIMF 20.00 %（1／5）;1例HES患者为野生型。PV、ET和CIMF突变患者拷贝数分别为2.14×102～1.5×107、9.80×102～4.4×107和4.10×103～3.70×106。结论　实时定量PCR检测JAK2 V617F基因简便、快捷, 并且易于定量, 检测阳性率与国外报道相似, 适合临床用于CMPD的诊断和疗效评估。     

隐匿性真性红细胞增多症一例并文献复习

目的:探讨隐匿性真性红细胞增多症的临床特征及诊断。方法:回顾性分析宁夏回族自治区人民医院收治的1例JAK2 V617F基因突变的隐匿性真性红细胞增多症患者的临床特点及诊疗经过,并进行相关文献复习。结果:该患者结合骨髓三系增生明显活跃,巨核细胞明显增多,且可见发育异常、多形性变及集簇,局灶纤维组织增生,JAK2 V617F基因突变检测阳性,全血细胞正常,诊断为隐匿性真性红细胞增多症,给予口服抗凝治疗后,患者病情稳定。结论:隐匿性真性红细胞增多症临床少见,临床特征和疾病演变需进一步观察研究。     

骨髓增殖性肿瘤JAK2V617F基因突变与疾病临床特征相关性的分析※

 摘要：目的：研究我区骨髓增殖性肿瘤（MPN）中JAK2V617F基因突变与疾病临床特征的相关性。方法：回顾性分析我区134例MPN患者JAK2V617F基因突变与患者性别、年龄、民族、外周血白细胞、血小板计数、血红蛋白、血清乳酸脱氢酶（LDH）水平及是否合并血栓、出血及心血管疾病并发症等临床特征的相关性进行统计分析。结果：134例MPN患者中PV 51例,ET 66例,IMF17例,存在JAK2V617F突变98例（73.1%）。年龄60岁以上者发生突变率较60岁以下者明显增高（p＜0.05）,MPD患者中JAK2V617F突变阳性者外周血白细胞计数、血红蛋白水平与血栓、心血管疾病的发生率均高于突变阴性者（p＜0.05）,而性别、民族、血小板计数及LDH水平在JAK2V617F基因突变阳性与阴性患者间无显著性差异（p＞0.05）。结论：MPN患者JAK2V617F基因突变与MPN患者年龄、外周血白细胞计数、血红蛋白水平、血栓及心血管疾病并发症相关。     

WHO bone marrow features and European clinical, molecular, and pathological (ECMP) criteria for the diagnosis of myeloproliferative disorders

The bone marrow criteria defined by the World Health Organization (WHO) are based on characteristic increase and clustering of morphologically abnormal enlarged megakaryocytes as a pathognomonic clue to describe three distinct phenotypic entities of myeloproliferative disorders (MPDs): (1) essential thrombocythemia (ET), (2) early and overt polycythemia vera (PV) and (3) prefibrotic, early fibrotic, and fibrotic chronic idiopathic myelofibrosis (CIMF-0, 1, 2 and 3). Based on established WHO bone marrow features, and the use of new molecular and laboratory markers including JAK2(V617F) mutation, endogenous erythroid colony (EEC) formation and serum erythropoietin (EPO), we present updated European clinical, molecular and pathological (ECMP) criteria for the differential diagnosis of true ET, PV and CIMF. As compared to the WHO bone marrow features, each of the laboratory and molecular markers are not sensitive enough for the diagnosis and classification of the three prefibrotic MPDs. The proposed WHO/ECMP criteria reduce the platelet count to the upper limit of normal (>400x10(9)l(-1)) as inclusion criterion for the diagnosis of thrombocythemia in true ET, early stages of PV and prefibrotic CIMF. The combined use of WHO and ECMP criteria differentiate PV from congenital and acquired erythrocytosis, true ET from reactive thrombocytosis and separates true ET from CIMF-0/1 mimicking ET. Only half of the patients with true ET and CIMF carry the JAK2(V617F) mutation (sensitivity 50%). Early PV mimicking ET is featured by the presence of JAK2(V617F) mutation, EEC, low serum EPO levels, normal hematocrit, and increased bone marrow cellularity due to increased erythropoiesis ("forme fruste" PV) when WHO/ECMP criteria are applied. The combination of JAK2(V617F) PCR test and increased hematocrit is diagnostic for PV (sensitivity 95%, specificity 100%). The degree of JAK2(V617F) positivity of granulocytes is related to disease stage: heterozygous in true ET and early PV and mixed hetero/homozygous to homozygous in overt and advanced PV and CIMF. Bone marrow histology assessment should remain the gold standard criterion for the diagnosis and staging of the MPDs true ET, PV and CIMF and its differentiation from primary or secondary erythrocytosis, reactive thrombocytosis and thrombocythemias associated with atypical MPD, myelodysplastic syndromes, and chronic myeloid leukemia,. The proposed WHO/ECMP criteria allow a cross talk between clinicians, pathologists and scientists to much better characterize the nature and natural history of each of the WHO/CMP defined early and overt MPDs.     

Usefulness of JAK2V617F mutation in distinguishing idiopathic erythrocytosis from polycythemia vera

Idiopathic erythrocytosis (IE) is a primary erythrocytosis not fulfilling the criteria for polycythemia vera (PV) diagnosis. In order to verify the relationship between IE and PV, we screened JAK2V617F mutation in a consecutive series of 11 IE and, for comparison, in 15 PV. JAK2V617F mutation was screened by both cDNA sequencing and mutation specific PCR in both peripheral blood and bone marrow samples. All 11 IE tested negative for JAK2V617F mutation, which, conversely, occurred in 11/15 (73.3%) PV. Our results demonstrate that JAK2V617F is absent in IE and may represent a useful molecular marker for distinguishing IE from PV.     

Diagnostic Performance of Erythropoietin Levels in Polycythemia Vera: Experience at a Comprehensive Cancer Center

IntroductionConsidering the evolving diagnostic criteria of polycythemia vera (PV), we analyzed the utility of serum erythropoietin (EPO) as a predictive marker for differentiating polycythemia vera (PV) from other etiologies of erythrocytosis.Patients and MethodsWe conducted a retrospective study after a review of electronical medical records from January 2005 to December 2016 with diagnosis of erythrocytosis using International Classification of Disease–specific codes. To evaluate the diagnostic performance of EPO levels and JAK2-V617F mutation, we constructed a receiver-operated characteristic curve of sensitivity versus 1-specificity for serum EPO levels and JAK2-V617F mutation as predictive markers for differentiating PV from other causes of erythrocytosis.ResultsWe surveyed 577 patients with erythrocytosis. Median patient age was 59.2 years, 57.72% (n = 329) were male, 86.3% (n = 491) were white, and only 3.3% (n = 19) were African American. A total of 80.88% (n = 351) of those diagnosed with PV had a JAK2-V617F mutation compared to only 1.47% (n = 2) whose primary diagnosis was secondary polycythemia. When comparing JAK2-V617 mutation to the EPO level, the area under the curve of JAK2-V617 (0.8970) was statistically larger than that of EPO test (0.6765). Therefore, the PV diagnostic methodology using JAK2-V617 is better than the EPO test. An EPO level of < 2 mIU/mL was > 99% specific to predict PV but was only 12% sensitive.ConclusionIn the appropriate clinical setting, cytogenetic and molecular studies such as JAK2 mutation status prevail as the most useful tools for PV case identification. The use of isolated EPO to screen patients with erythrocytosis is not a good diagnostic approach.     

Somatic JAK-2 V617F Mutational Analysis in Polycythemia Rubra Vera: a Tertiary Care Center Experience

Background: Polycythemia rubra vera (PV), being a primary polycythemia, is caused by neoplastic proliferation of erythroid, megakaryocytic and granulocytic lineages which result in panmyelosis. PV patients have a somatic acquired mutation in the Janus kinase (JAK2) pathway, rendering cell proliferation independent of the normal regulatory mechanisms that regulate erythropoiesis. The rational of this study was to determine the prevalence of the JAK-2 V617F mutation in Pakistani patients with PV. Materials and Methods: In this cross sectional study, 26 patients with PV were enrolled from January 2010 to December 2014. Patients were diagnosed based on WHO criteria for PV. All were screened for G-T point mutation (V617F) in the JAK2 gene on chromosome 9 by an allele specific PCR. Results: The mean age was 53.49.31 years (range 36-72) and the male to female ratio was 2:1. The frequency of JAK2 V617F positivity in our PV patients was found to be 92.3%. Overall 30.7% of patients were asymptomatic and remaining 69.3% presented with symptomatic disease. The mean hemoglobin was 18.11.9g/dl with the mean hematocrit of 55.68.3%. The mean total leukocyte count was 12.87.1x109/l and the platelet count was 511341.9x109/l. A positive correlation of JAK2 V617F mutation was established with high TLC count (P=0.01). No correlation of JAK2 V617F could be established with age or gender (P>0.05). Conclusions: The JAK2 V617F mutation frequency in our PV patients was similar to those reported internationally. Screening for the mutation in all suspected PV cases could be beneficial in differentiating patients with reactive and clonal erythrocytosis.     

JAK2V617F mutation and spontaneous megakaryocytic or erythroid colony formation in patients with essential thrombocythaemia (ET) or polycythaemia vera (PV)

The in vitro cultures of erythroid (BFU-E) and megakaryocytic (CFU-Meg) progenitors have been useful diagnostic tools in myeloproliferative disorders (MPD). However, after the discovery of the JAK2V617F mutation, their diagnostic role has been uncertain. In this single-centre retrospective study we analyzed JAK2V617F mutation in 58 ET and 42 PV patients diagnosed according to WHO criteria and compared the results with those of colony forming assays with special emphasis on CFU-Meg growth. 91% of PV and 57% of ET patients had JAK2V617F mutation and they all showed spontaneous BFU-E growth. However, endogenous BFU-E formation was also seen in nine JAK2V617F mutation negative patients displaying also a normal JAK2 exon 12 allele. Endogeneous CFU-Meg colony formation was found in 59% of PV and 53% of the ET patients. A subgroup of ET patients (n=7) displayed sole spontaneous CFU-Meg growth without spontaneous BFU-E growth. They all were JAK2 mutation negative, but one of them had MPL mutation. In conclusion, in vitro cultures of haematopoietic progenitors are sensitive diagnostic tools in the present group of 100 MPD patients revealing also JAK2 mutation negative ET and PV patients displaying sole spontaneous CFU-Meg or BFU-E growth.     

Prevalence and clinicopathologic correlates of JAK2 exon 12 mutations in JAK2V617F-negative polycythemia vera.

After accounting for misdiagnosis and treatment effect, allele-specific (AS)-PCR detects the JAK2V617F mutation in >95% of polycythemia vera (PV) patients. Using database inquiry, we identified 6 of a total 220 cases with PV that were JAK2V617F-negative (prevalence=3%). Of these, five cases ( approximately 80%) were found to harbor one of the two JAK2 exon 12 mutations (F537-K539delinsL or N542-E543del) in bone marrow (BM) and/or peripheral blood cells. Similar screening of six additional cases - three each with idiopathic erythrocytosis (IE) or otherwise unexplained erythrocytosis (UE) - did not reveal either JAK2V617F or JAK2 exon 12 mutations. We found JAK2 exon 12 mutations in PV cases to be readily detected by both DNA sequencing and AS-PCR, regardless of whether BM or peripheral blood cells were used as the source for DNA. Although erythroid hyperplasia was the predominant histologic feature on BM examination, megakaryocyte abnormalities and reticulin fibrosis were noted in most PV patients harboring exon 12 mutations. However, similar BM morphologic changes can also be seen in some JAK2V617F-positive PV cases; therefore, distinct genotype-phenotype association cannot be established.     

Coexisting JAK2V617F and CALR Exon 9 Mutations in Myeloproliferative Neoplasms - Do They Designate a New Subtype?

The classic BCR-ABL1-negative myeloproliferative neoplasm is an operational sub-category of MPNs that includes polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF). The JAK2V617F mutation is found in ~ 95% of PV and 50-60% of ET or PMF. In most of the remaining JAK2V617F- negative PV cases, JAK2 exon 12 mutations are present. Amongst the JAK2V617F-negative ET or PMF 5-10% of patients carry mutations in the MPL gene. Prior to 2013, there was no specific molecular marker described in the remaining 30-40% ET and PMF. In December 2013, two research groups independently reported mutations in the gene CALR found specifically in ET (67-71%) and PMF (56-88%) but not in PV. Initially CALR mutations were reported mutually exclusive with JAK2 or MPL. However, co-occurrence of CALR mutations with JAK2V617F has been reported recently in a few MPN cases. Many studies have reported important diagnostic and prognostic significance of CALR mutations in ET and PMF patients and CALR mutation screening has been proposed to be incorporated into WHO diagnostic criteria for MPN. It is suggestive in diagnostic workup of MPN that CALR mutations should not be studied in MPN patients who carry JAK2 or MPL mutations. However JAK2V617F and CALR positive patients might have a different phenotype and clinical course, distinct from the JAK2-positive or CALR-positive subgroups and identification of the true frequency of these patients may be an important factor for defining the prognosis, risk factors and outcomes for MPN patients.