MYH9综合征的免疫荧光检测与基因分析

Immunofluorescence Method and Gene Analysis for Diagnosis of MYH9 Syndrome

目的研究两个MYH9综合征家系中性粒细胞包涵体的本质及基因突变。方法瑞氏染色观察先证者及其家系患者的外周血片血小板和中性粒细胞的特殊形态。间接免疫荧光方法结合DAPI复染技术观察正常对照和患者血片非肌性肌球蛋白IIA的亚细胞定位。从先证者及家系成员外周血中白细胞中提取基因组DNA,PCR扩增MYH9的40个外显子及侧翼内含子序列,检测其基因突变。结果患者外周血片中性粒细胞胞浆中可见明显团块状、半月形的绿色荧光聚集,其大小、位置与瑞氏染色的中性粒细胞包涵体相同。Fechtner综合征MYH9基因改变为外显子40第5981位核苷酸C→T杂合改变,使第1933位密码子(CGA,编码Arg)突变为终止密码TGA。May-Hegglin异常MYH9基因改变为外显子38第5701位核苷酸G→A杂合改变,使1841位谷氨酰氨变为赖氨酸。结论建立了诊断MYH9综合征的一种独特的免疫荧光方法。R1933X和E1841K杂合改变是导致Fechtner综合征和May-Hegglin异常的原因。

Objective To identify the property of the inclusions in neutrophils and gene mutation of MYH9 syndrome in two chinese families. Methods Indirect immunofluorescence with DAPI staining technology was used to observe the location of nonmuscle myosin IIA in patients＇ peripheral blood smear. All the exons and exon-intron boundaries of the MYH9 gene were amplified by PCR followed by direct sequencing. Results There appears abnormal nonmuscle myosin IIA aggregates in cytoplasm of neutuophils which is similar to the Wright＇s-Giemsa staining in terms of location and size of inclusions. A heterozygous C to T mutation and heterozygous G to A mutation Were found in Fechtner syndrome family at nucleotide 5981 and in May-Hegglin anomaly family at nucleotide 5701 respectively. Conclusion Immunofluorescence for nonmuscel myosin IIA is a simple and sensitive method for the diagnosis of MYH9 syndrome. R1933X and E1841K are causative genetic defects of Fechtner syndrome and May-Hegglin abnomally.