急性淋巴细胞白血病中T细胞受体δ基因Vδ2－Dδ3及Dδ2－Dδ3重排结合部顺序的研究

Junctional sequences of T cell receptor Vδ2-Dδ3 orDδ2-Dδ3 rearrangements in acute lymphoblasticIeukemia

为了探讨中国人Ｔ细胞受体（ＴＣＲ）δ基因的Ｖ－Ｄ－Ｊ结合部顺序（Ｎ顺序）的组成，及其在急性淋巴细胞白血病（ＡＬＬ）患者中的重组规律，应用聚合酶链反应（ＰＣＲ）对４６例ＡＬＬ标本进行了ＴＣＲδ基因重排的研究，发现１６例患者出现Ｔ细胞受体δ基因不完全重排，其中１３例为Ｖδ２－（Ｄ）－Ｄδ３重排，３例为Ｖδ２－（Ｄ）－Ｄδ３合并Ｄδ２－Ｄδ３重排。进而应用ＤＮＡ直接测序技术，对这１６例ＡＬＬ标本ＴＣＲδ基因重排结合部进行了分析，证实由于Ｖδ２或Ｄδ２的３＇端和Ｄδ３的５＇端碱基缺失、部分Ｄδ１或Ｄδ２顺序存在、Ｎ顺序的插入以及未发生缺失的Ｖδ２，Ｄδ２或Ｄδ３编码顺序端Ｐ核昔酸的非随机插入，使得每例患者的结合部顺序均存在高度个体特异性。此外，对Ｎ顺序碱基成份的分析表明，其脱氧鸟嘌呤和脱氧胞嘧啶含量大于７０％，而脱氧腺嘌呤和脱氧胸腺嘧啶小于３０％，说明碱基插入并非完全随机。本文结果提示ＴＣＲδ基因重排是淋巴细胞发育中的一个早期事件，其结合部顺序的测定，可作为检测ＡＬＬ患者微小残留病变的特异性克隆标志。

-cell receptor (TCR) δ chain gene rearrange-ments were studied by polymerase chain reactiontPCRi analysis in 46 patients with acute lymphoblasticleukemia (ALL). Sixteen patients were found to haveincomplete rearrangements of the TCR δ genes. A-mong them, 13 patients displayed Vδ2-Dδ3 rearrange-ment. while 3 had both Vδ2-Dδ3 and Dδ2-Dδ3 rear-rangements. To determine the junctional sequence ofTCRδ gene, PCR products from the 16 patients weresequenced directly or after M13 cloning. The resultsshowed the. junctional sequences of TCRδ gene arehighly specific for each allele. This sequence diversityresulted from several factors including deletion of the 3’end of Vδ2 or Dδ2 segment and 5’ end of Dδ3 seg-ment , the presence of Dδl or Dδ2 sequences, insertionof N nucleotides and the association of P nucleotideswith intact Vδ2 and Dδ3 segments. In addition , analy-sis of Nnucleotide contents revealed that the amount ofGC was much larger than that of AT (70% : 30%) ,indicating the insertion of N nucleotide was not fullyrandom. Our sequence data confirmed that the imcom-plete rearrangement of TCRδ gene is an early event inthe lymphoid cell ontogenesis, and its N sequences inV-(D)J junctional region may be used as a specificmarker of clonality to detect the minimal residual dis-ease (MRD) in ALL.