ABNORMALITIES OF PIG-A TRANSCRIPTS IN GRANULOCYTES FROM PATIENTS WITH PAROXYSMAL-NOCTURNAL HEMOGLOBINURIA
ABNORMALITIES OF PIG-A TRANSCRIPTS IN GRANULOCYTES FROM PATIENTS WITH PAROXYSMAL-NOCTURNAL HEMOGLOBINURIA
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DOI：
10.1056/nejm199401273300404
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发表时间：
1994-01-27
影响因子：
158.5
通讯作者：
KINOSHITA, T
中科院分区：
文献类型：
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作者：
MIYATA, T;YAMADA, N;KINOSHITA, T
背景。阵发性睡眠性血红蛋白尿 (PNH) 是一种后天性疾病，造血细胞合成将蛋白质锚定在细胞膜上的糖基磷脂酰肌醇分子存在缺陷。最近，我们证明了一种名为 PIG-A（A 类磷脂酰肌醇聚糖）的基因（糖基磷脂酰肌醇生物合成的一个组成部分）与两名患者的 PNH 有关。本研究旨在阐明 PIG-A 是否是导致所有 PNH 病例的基因，并进一步表征该基因的体细胞获得性异常。方法。我们研究了 15 名 PNH 患者的粒细胞。通过荧光激活流式细胞术评估CD55和CD59的细胞含量。 PIG-A 转录物被逆转录，通过聚合酶链式反应扩增，并克隆到质粒中。通过核苷酸测序分析克隆的互补DNA的结构，并根据转染后其将PIG-A缺陷细胞系的异常表型恢复正常的能力来评估其功能。结果。三名患者的 PIG-A 转录物大小异常，且模式不同，其中一名患者的 PIG-A 转录物水平非常低。 11 名患者的转录本大小正常，但转染检测显示，每位患者中的一些转录本没有功能。非功能性 PIG-A 转录物的百分比与受影响的粒细胞的百分比相关 (P
Background. Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired disorder in which there is a deficiency in the synthesis by hematopoietic cells of the glycosyl-phosphatidylinositol molecules that anchor proteins to the cell membrane. Recently, we demonstrated that a gene termed PIG-A (for phosphatidylinositol glycan class A), a component of glycosyl-phosphatidylinositol biosynthesis, was responsible for PNH in two patients. The present study was undertaken to elucidate whether PIG-A is the gene responsible for all cases of PNH and to characterize further the somatically acquired abnormalities of this gene.Methods. We studied granulocytes from 15 patients with PNH. The cell content of CD55 and CD59 was assessed by fluorescence-activated flow cytometry. PIG-A transcripts were reverse-transcribed, amplified by the polymerase chain reaction, and cloned into plasmids. The structure of the cloned complementary DNA was analyzed by nucleotide sequencing, and its function was assessed on the basis of its ability to restore to normal the abnormal phenotype of a PIG-A-deficient cell line after transfection.Results. Three patients had size abnormalities of PIG-A transcripts with different patterns, and in one patient a very low level of the PIG-A transcript was found. Eleven patients had transcripts of normal size, but the transfection assay revealed that in each patient some of them were nonfunctional. The percentage of nonfunctional PIG-A transcripts was correlated with the percentage of affected granulocytes (P