【作者】 武永强；

【导师】 刘延方；

【作者基本信息】 郑州大学 ， 内科学， 2012， 硕士

【摘要】 背景和目的在急性白血病的诊断中,细胞形态学联合细胞化学将急性白血病分型的准确性达89%,白血病免疫分型的准确性及灵敏度都较高,能够将急性白血病的分型诊断准确率大大提高,其在急性白血病的现代诊断中起着非常重要的作用。急性白血病的免疫分型是基于不同的急性白血病细胞表达有其特定的细胞表面抗原群,应用抗原抗体特异性结合的原理,借助于流式细胞仪,对急性白血病细胞膜表面表达的抗原进行分型,能够对那些形态学不典型,仅靠细胞形态和化学染色难以诊断的病例做出精确诊断,尤其是对急性淋巴细胞白血病(acute lymphoblastic leukemis, ALL)的诊断,不仅能够将ALL与与急性髓系白血病(acute myeloid leukemis, AML)区别开,更能将B细胞ALL与T细胞ALL进行区分,弥补了ALL形态学诊断的不足。但白血病免疫分型检测中发现有些AML细胞不仅表达髓细胞相关抗原,而且还表达淋巴细胞相关抗原,反之亦然。这种情况是伴系抗原表达,还是混合型白血病,较难确诊。因此选择更加特异的标志物以明确诊断非常必要,这对病人的治疗、预后具有重要的作用。我们在常规免疫分型检测的基础上,加做胞浆抗原染色,以探讨白细胞分化特异性胞浆抗原在急性白血病细胞的表达及意义。方法选择2010年9月至2011年5月在我院血液科住院治疗的急性白血病患者138例,男86例,女52例,年龄14岁-76岁,中位年龄37.5岁。所有患者经血常规、生化、骨髓细胞形态学及化学染色、白血病免疫分型及细胞遗传学诊断。应用流式细胞术三色荧光标记,对细胞膜抗原进行常规检测,对形态学分类难以诊断,出现跨系列表达的加测胞浆内抗原检测。髓系选择胞浆抗原cMPO、急性B淋巴细胞白血病选择胞浆抗原cCD79a、急性T淋巴细胞白血病选择胞浆抗原cCD3。结果1.诊断按FAB分型138例AL中诊断为AML者76例,ALL者49例,不能明确分型者13例。应用流式细胞术进行白血病免疫分型分析,ALL49例包括B-ALL40例和T-ALL9例。13例根据细胞形态无法确诊的病例最终诊断2例AML-MO、5例B-ALL、2例T-ALL和4例混合细胞白血病。2.跨系抗原表达急性白血病中出现跨系抗原表达的占41.3%,其中AML、B-ALL和T-ALL中出现跨系抗原表达的比例分别为50%、26.7%、18.1%。AML中跨系抗原表达以CD7(53.8%)和CD19(30.8%)为主。出现跨系抗原表达的B-ALL中CD33阳性率为83.3%,显著高于CD2及CD13(P<0.05)。3.敏感性和特异性本组所用抗体对AML的敏感性从高到低依次为CD33、CD13、cMPO、CD117,特异性依次为CD117、cMPO、CD13、CD33,胞浆抗原cMPO的敏感性低于CD33和CD13,但特异性高于CD33和CD13,cMPO和CD117的特异性无显著差别(P=-0.312),但cMPO敏感性高于CD117(P=-0.027)；本组所用抗体对B-ALL的敏感性依次为CD19、cCD79a、CD10、CD20,cCD79a特异性和CD10无显著差别(P=-0.312),和CD20无显著差别(P=0.287),但显著高于CD19(P=0.018)；本组抗体对T-ALL的敏感性从高到低依次为cCD3、CD7、CD2,特异性依次为cCD3、CD2、CD7,cCD3的敏感性高于CD7(P=0.042)和CD2(P=-0.047),特异性和CD2无差别(P=0.426),显著高于CD7(P=0.022)。4.胞浆抗原cMPO、cCD79a、cCD3的表达cMPO在AML、B-ALL和T-ALL中阳性率分别为87.5%、9.5%和0,cMPO在AML的表达率显著高于ALL(P<0.05),其中B-ALL与T-ALL之间的差别无显著性(P>0.05)；cCD79a在B-ALL、T-ALL和AML中的阳性率分别为92.7%、9.0%和0。cCD79a在B-ALL的表达率显著高于T-ALL(P<0.05),其中T-ALL与AML之间的差别无显著性(P>0.05)；cCD3在T-ALL中表达率为100%,在AML和B-ALL中均无表达。结论1.胞浆抗原cMPO、cCD79a、cCD3分别是AML、B-ALL、T-ALL的特异性标志,系列特异性高于细胞膜表面抗原。2.胞浆抗原cMPO、cCD79a、cCD3的检测有助于细胞形态不典型、细胞膜出现跨系抗原表达的白血病的准确分型及混合性白血病的正确诊断。更多还原

【Abstract】 Background and ObjectiveCell morphology and cytochemistry can improve the classification accuracy of acute leukemia up to89%in the diagnosis of acute leukemia. Because of the high accuracy and sensitivity, immunophenotyping plays an important role in the diagnosis of leukemia. The immunophenotype of acute leukemia is method to classify acute leukemia, based on the specific antigens expressed on acute leukemia blasts, which bind to specific antibodies conjugated with fluorescent detected by flow cytometer. This technique is used in the diagnosis and classification of acute leukemia cases with atypical morphology which cannot be diagnosed by bone marrow morphology and cytochemistry. In the diagnosis of acute lymphoblast leukemia, immunophenotype can be used to differentiate ALL from AML as well as to differentiate B-All from T-ALL, which is a compliment to the morphological diagnosis of ALL. However, it is found that AML leukemic cells express not only myeloid-associated antigen, but also the lymphocyte-associated antigen and vice versa. It is difficult to distinguish cross-lineage expression from mixed-lineage acute leukemia (MAL). Therefore it is necessary to choose more specific markers to confirm the diagnosis of acute leukemia, which plays an important role in the patient’s treatment and prognosis. On the base of the immunophenotype of cellular surface antigens, we performed a cytoplasmic antigen staining in order to investigate the expression of lineage specific cytoplasmic antigens in patients with acute leukemia and its significance.MethodsOne hundred and thirty-eight cases of acute leukemia were treated in the Department of Hematology of the First Affiliated Hospital of Zhengzhou University form September2010to May2011, including86male cases and52female cases. These patients range from14years old to76years old, with the median age at37.5years old.All patients were diagnosed by CBC, liver function, renal function, bone marrow cell morphology and histochemistry, immunophenotype and cytogenetics.The membrane antigens were detected in138patients with acute leukemia using three-color fluorescent by flow cytometry. Cytoplasmic antigens were detected in cases which difficulty to diagnose with morphological classification and cases with cross-lineage expression. Cytoplasmic antigens cMPO was selected for myeloid, Cytoplasmic antigens cCD79a was selected for acute B-lymphoblastic leukemia, and Cytoplasmic antigens cCD3was selected for acute T-lymphoblastic leukemia.Results1. DignosisAccording to the FAB classification,76out of138cases were diagnosed as AML and49/138cases were diagnosed as ALL, and other13/138cases were not explicitly classified. By immunophenotype, the49/138cases of ALL were subdivided into B-ALL (40cases) and T-ALL (9cases). In the cases morphologically not defined subtypes,2/13cases were diagnosed as AML-MO,5/13cases as B-ALL,2/13cases as T-ALL and4/13cases as mixed-lineage acute leukemia.2. The expression of cross-lineage antigensThere were41.3%of acute leukemia express Cross-lineage antigen. The cross-lineage antigen expression rates were50%in AML,26.7%in B-ALL and18.1%in T-ALL. Cross-lineage antigen expression in AML is mainly CD7(53.8%) and CD19(30.8%). CD33was expressed in83.3%of B-ALL, significantly higher than that of CD13and CD2(P<0.05).3. The specificity and sensitivityThe sensitivity of the antibody used in the diagnosis of AML were listed from high to low as CD33, CD33, cMPO, CD117, and the specificity from high to low as CD117, cMPO, CD13, CD33. The sensitivity of cytoplasmic antigens cMPO was lower than that of CD33and CD13, but the specific was higher than that CD33and CD13.There was no significant difference in the specificity between cMPO and CD117(P=0.312), but the sensitive of cMPO was higher than CD117(P=0.027). The sensitivity of the antibody used in the diagnosis of B-ALL were listed from high to low as CD19, cCD79a, CD10, CD20. There was no significant difference with specificity between cCD79a and CD10(P=0.312), CD20(P=0.287), but it was significantly higher than that of CD19(P=0.018). The sensitivity of antibodies used in the diagnosis of T-ALL were listed from high to low as cCD3, CD7, CD2, the specific from high to low as cCD3, CD2and CD7.The sensitive of cCD3was higher than CD7(P=0.042) and CD2(P=0.047), The specificity of cCD3had no difference with CD2(P=0.426), but it was significantly higher than that of CD7(P=0.022).4. The expression of Cytoplasmic antigen cMPO, cCD79a and cCD3The positive rates of cMPO were88.6%in AML,9.5%in B-ALL and0in T-ALL. The expression of cMPO in AML was significantly higher than that in ALL (P<0.05). There was no statistically significance in the expression of cMPO between B-ALL and T-ALL (P>0.05). The positive rates of cCD79a were92.7%in B-ALL,9.0%in T-ALL and0in AML. There was no statistically significance in the expression of cCD79a between T-ALL and AML (P>0.05). The positive rate of cCD3in T-ALL was100%. There were no cCD3expression in AML and B-ALL.Conclusion1. Cytoplasmic antigen cMPO, cCD79a, cCD3are specific markers of AML, B-ALL, and T-ALL. Their lineage specificity is higher than the cell surface antigen2. The detection of Cytoplasmic antigen cMPO, cCD79a, cCD3plays an importment role in the accurate diagnosis and classification of acute leukemia, especially in the cases with atypical morphology and cross-lineage antigen expression, such as mixed-lineage acute leukemia.更多还原