一例X-连锁高IgM综合征合并进行性多灶性脑白质病的基因变异分析

目的对1例临床拟诊为X-连锁高IgM综合征(X-linked hyper-IgM syndrome,XHIM)并发进行性多灶性脑白质病变(progressive multifocal leukoencephalopathy,PML)的患儿CD40L基因及人类嗜神经多瘤病毒(Jamestown Canyon virus,JCV)进行检测,明确致病原因,为临床诊断提供依据。方法采集患儿及父母外周血提取基因组NDA,设计扩增CD40L基因5个外显子及外显子-内含子连接区的特异性引物并进行PCR扩增,产物测序结果与GenBank中CD40L基因序列分析对比确定有无变异。用两对JCV特异性引物对患儿外周血DNA行巢式PCR扩增,目的条带PCR产物测序结果与GenBank中JCV序列对比确定患儿是否存在JCV感染。结果测序结果显示患儿为CD40L c.506 A>C(p.Tyr169Ser)错义变异半合子,该变异位于CD40L肿瘤坏死因子同源区结构域,引起CD40L蛋白疏水作用及结构稳定性丧失,经PolyPhen2及SIFT蛋白功能预测软件预测分别为很可能有害变异(probably damaging,score=1.00)及有害变异(deleterious,score=-8.868),该变异尚未见文献报道。患儿母亲携带CD40L c.506 A>C(p.Tyr169Ser)半合子变异,父亲未检测到该变异。患儿外周血DNA巢式PCR产物经凝胶电泳后发现JCV目的条带,测序结果与GenBank中JCV基因序列比对同源性达到99%,表明患儿外周血DNA中含有JCV,有JCV感染。结论CD40L基因分析及JCV检测结果确诊患儿为XHIGM,其并发的PML与JCV感染有关。

Genetic analysis of a child with co-commitment progressive multifocal leukoencephalopathy and X-linked hyper IgM syndrome

Objective To detect variant of the CD40L gene and infection of Jamestown Canyon virus(JCV)in a 7-year-and-9-month-old boy with co-commitment progressive multifocal leukoencephalopathy(PML)and X-linked hyper IgM syndrome(XHIGM).Methods Peripheral blood samples of the child and his parents were collected for the extraction of genomic DNA.The 5 exons and exon/intronic boundaries of the CD40L gene were subjected to PCR amplification and sequencing.Suspected variants were analyzed by using bioinformatic software.The JCV gene was amplified from genomic DNA by nested PCR and sequenced.Results The child was found to harbor a hemizygous c.506 A>C(p.Y169S)missense variant in exon 5 of the CD40L gene.The variant may affect the TNFH domain of the CD40L protein and result in structural instability and loss of hydrophobic interaction between CD40L and CD40.As predicted by PolyPhen2 and SIFT software,the variant was probably damaging(score=1.00)and deleterious(score=-8.868).His mother was found to be a heterozygous carrier,while the same variant was not found in his father.Gel electrophoresis of the nested PCR product revealed presence of target JCV band,which was confirmed to be 99%identical with the JCV gene by sequencing.Conclusion The patient was diagnosed with co-commitment XHIGM and PML based on the testing of the CD40L gene and JCV infection.