【作者】 刘丹丹；

【导师】 陈子兴；

【作者基本信息】 苏州大学 ， 内科学， 2009， 博士

【摘要】 一、MDS患者的FAB分型与WHO分型的临床、实验室特点为了探讨骨髓增生异常综合征(Myelodysplastic syndrome,MDS)患者的FAB分型及WHO分型的形态学、细胞遗传学和预后特点以及不同亚型的实用意义,将440例FAB分型(RA、RAEB亚型)及重新分类后的264例WHO分型的MDS患者的临床(年龄、性别及预后)及实验室(外周血、骨髓、染色体及IPSS风险分组)特点进行总结分析。发现MDS患者的FAB分型及WHO分型各个亚型的上述特点各有不同;这两种亚型相比较,WHO分型在与临床预后的相关性方面稍弱于FAB分型,但WHO分型更加细化了FAB分型,将MDS的诊断分型与其临床及实验室特征更为紧密的结合起来,减少了亚型内的异质性。因此,对于国内MDS患者来讲,可根据临床及科研的不同需要选择不同的分型。而国内的MDS患者的临床及实验室特点与国外MDS患者相比具有异同点,说明由于民族、种族及地域的差别,MDS患者的特点不尽相同。二、核型异常MDS-RA的细胞形态学、细胞遗传学特点及两者的关系探讨骨髓细胞病态造血是MDS的特点,也是MDS-RA亚型的主要诊断依据。但是,关于MDS-RA患者骨髓细胞形态学异常变化的程度、类型及频率尚没有“量化”的研究报道;细胞形态变化的特点与染色体异常的统计学关系更为罕见。因此,为了研究MDS患者的骨髓细胞形态变化的大体“图像”,使我们更深入的了解MDS患者的形态学变化的特点,为MDS的诊断提供更坚实、更丰富的形态学理论依据。本研究以207例非MDS贫血及46例正常核型MDS患者为对照,对48例核型异常MDS-RA患者的形态学特点、细胞遗传学特点及二者的关系进行分析探讨。发现核型异常的MDS-RA患者的骨髓细胞形态学异常发生率较正常核型MDS及非MDS贫血患者高,骨髓中病态表现的类型较多,病态细胞比例较高;而某些形态学改变的发生及病态细胞的数量与染色体异常的方式及程度有关。因此研究核型异常RA的形态学及细胞遗传学改变的特点有助于对MDS-RA的诊断及鉴别诊断。三、骨髓增生异常综合征的骨髓细胞免疫表型特点近年来,流式细胞术免疫表型的研究及应用越来越广泛,人们从不同的角度及方面对MDS的免疫标志进行了探讨,其中研究最多的为MDS的原始细胞的表型特点。但是MDS患者骨髓幼稚细胞群体的表型特征如何?常规的简单的免疫标志对于MDS患者的诊断及鉴别诊断的意义又如何呢?因此,通过应用直接免疫荧光法采用流式细胞仪对36例MDS,18例从和11例正常人的免疫表型进行检测,发现FAB各亚型的免疫表型特点不同;RAEB的患者其CD13、CD33、CD34的表达占优势;MDS高危组患者与正常人比较CD19、CD20的表达降低,而CD13、CD33的表达偏高(P＜0.05);MDS患者与AA患者在T、B淋巴细胞系及髓细胞系的免疫标志表达方面均有显著性差异。因此,①应用常规免疫表型检测骨髓单个核细胞有助于鉴别诊断AA与MDS,预测低危MDS向高危MDS转化。②MDS患者的髓系抗原的表达占优势,而B淋系抗原表达有降低趋势。四、DNTT及BLNK基因在MDS患者的表达MDS的发病机制仍不明确,是一个涉及多基因、多因素、多步骤的异质性疾病。而基因芯片可以短时间内对成千上万个基因进行高通量的检测。通过本研究前期MDS患者CD34+细胞基因芯片的检测,发现与B淋巴细胞相关的BLNK基因及DNTT基因有下调趋势。为了探讨MDS发生的可能机制及分子生物学的特点,进一步通过对41例MDS患者、35例非MDS贫血、4例CAA、6例AML患者及22例正常人骨髓单个核细胞的实时定量PCR检查,发现:①MDS患者DNTT基因的表达与正常对照、非MDS贫血患者比较,表达降低,统计学有显著性差异(P＜0.05)。②RAEB与RA比较,BLNK基因表达减低,具有统计学意义(P＜0.05)。③MDS患者BLNK基因的表达与DNTT基因的表达存在正相关关系。因此,MDS患者的DNTT基因及BLNK基因表达下调,可能是引起MDS患者B淋系抗原减低的原因之一;DNTT及BLNK基因的检测也许有助于MDS的诊断及鉴别诊断。更多还原

【Abstract】 1 The clinic and laboratory features of MDS patients by FAB and WHO classificationTo study the features of morphological,cytogenetic and prognosis of MDS patients in FAB and WHO classification and difference of clinic useful value between the classifications,the clinic(age,gender and prognosis) and laboratory(peripheral blood, bone marrow’chromosome and IPSS groups) features in 440 MDS patients of FAB subtypes and 264 patients reclassified by WHO subtype were summarized and analyzed. The difference of these features in each subtypes of two classifications were showed respectively.Although the correlations between subtype and prognosis in WHO-classification were no more tightly than that in FAB-classification,the former was more elaborate than the latter when compared the two classifications.WHO classification reduced the heterogeneity because each subtype of it closely correlated with the features of clinic and laboratory.Therefore,different classification can be chosed by different demand in clinical work and scientific research for our Chinese patients of MDS.The different features in MDS between China and other countries indicate that the influence factors of race,country and region have impact on the clinic and laboratory features in MDS patients.2 The morphological and cytogenetic features and theirs correlations in abnormal karyotype MDS-RA patientsDysplasia is not only the important characteritics,but also the necessary evidence in MDS.But to date,there was no "quantization" study reported about the degree,frequency and types of dysplasias in MDS-RA.Analysis on the correlation between morphology and cytogenetics was hardly found before.Therefore,to study the ’whole picture’of dysplasias of bone marrow cells in MDS,comprehend the dysplastic characteristics deeply,provide the substantial and copious theoretical evidence in morphology,48 RA patients with abnormal karyotype were analyzed for the features of morphological,cytogenetical abnormalities and the relationships between them.46 normal karyotype MDS-RA and 207 non-MDS anemia patients were enrolled as control groups.Morphological abnormalities more frequently happened in abnormal karyotype MDS-RA patients than that of the patients in other groups;more diverse types and higher proportions of dysplasia cells in bone marrow were found in RA patients with abnormal karyotypes;The frequency of some special dysplasia and number of some dysplastic cells may be influenced by the chromosome abnormalities.The study on the features of morphology and cytogenetics in abnormal karyotype MDS-RA patients are helpful for the diagnosis of MDS.3 The immunophenotypic features of bone marrow cells in MDSFlow cytometry was widely used on the detection and study of immunophenotypes in recent years.Immunophenotype of MDS was explored from different aspect and purpose, and most of studies were about the characteristics of the blast phenotype.But what are the immunophenotypic features of immature cells in the bone marrow? How about the significance on the application with conventional and simple immunological markers in MDS patients? And what value about it in the disgnosis of MDS? Using flow cytometer with direct immunofluorescence methods,immunophenotypes of 36 MDS patients(in FAB classification),18 AA patients and 11 healthy people were determined.Then R banding technique was used to perform chromosome karyotype analysis.Different immunophenotypic features was detected in each subtypes;expression of CD13、CD33 and CD34 was predominant in RAEB patients;expression of CD19、CD20 decreased and expression of CD13、CD33 increased in RAEB/RAEBT patients compared with those in healthy persons(P＜0.05).Statistically significant difference on the T,B lymphoid antigens and myeloid markers were found between MDS patients and AA patients.Thus, conventional flow cytometry detecting the immunophenotype of bone marrow cells are helpful in the differential diagnosis between MDS and AA,predict the progress from lower risk to higher risk subtypes.The expression of myeloid antigens was predominant and the expression of B lymphoid markers showed a decreased expression in MDS patients.4 Expression of genes DNTT and BLNK in patients with MDS MDS is a heterogenic disease that involves multiple genes,factors and steps.Its pathogenesis is not clear yet.Gene chip technique is a method that can be used to the high-throughout examination of thousands of genes in a short period.The previous study of this project in which the gene chip was used to detect the CD34+ cell of MDS patients revealed that a down-regulated tendency in the expression of gene DNTT and gene BLNK which were related to B-lymphocytes.Therefore,to investigate the possible pathogenesis and molecular characteristics of MDS,a real-time PCR experiment was performed next step in 41 MDS,35 non-MDS anemia,4 CAA,6 AML patients and 22 healthy controls. The experimental results showed that:①compared with the normal controls and non-MDS anemia patients,the MDS patients had a decreased expression of DNTT with a statistical significance(P＜0.05);②compared with that in RA patients,BLNK gene had a decreased expression in RAEB patients with statistical significance(P＜0.05);③the expression of BLNK gene in MDS had a positive correlation with the expression of DNTT gene.Therefore,the down-regulated expression of DNTT and BLNK gene might be one cause of the reduction of B lymphoid antigens in MDS;the detection of DNTT and BLNK gene may be helpful for the diagnosis of MDS.更多还原