急性B淋巴细胞性白血病患儿外周血T淋巴细胞克隆谱系分析

Analysis of T cell repertoire in children with acute B lymphoblastic leukemia

目的 观察儿童急性B淋巴细胞性白血病化疗前后T淋巴细胞克隆谱系的异常。方法 逆转录 聚合酶链反应 (RT PCR)及高压变性聚丙烯酰胺凝胶电泳法检测 8例正常对照组小儿及 12例急性B淋巴细胞性白血病患儿化疗前外周血T细胞受体 (TCR) β链可变区 (BV)第三互补决定区(CDR3)的克隆谱系 ,4例患儿化疗后重新取材。结果 (1) 12例患儿化疗前BV2和BV3表达较正常对照组增高 (P均 <0 0 5 ) ,BV17和BV18表达降低 (P均 <0 0 5 )。 4例患儿治疗前BV2 1家族低表达 ,缓解后表达进一步降低 (P <0 0 5 )。 (2 )正常对照组TCRBV多数家族CDR3谱型为正态分布 ,12例患儿TCRBVCDR3谱型异常发生率为 14 % ,高于正常对照组的 5 5 % (P <0 0 5 ) ,发生谱型异常较多的家族依次是BV14、BV1、BV16、BV2 0、BV13 1、BV13 2和BV6。 4例患儿第 1次化疗缓解后 3个月谱型异常的家族均恢复正态分布。结论 急性B淋巴细胞性白血病患儿部分T细胞克隆发生异常增生或缺失 ,提示白血病患儿细胞免疫功能异常 ,化疗初次缓解后 3个月T细胞克隆谱系基本恢复正常的正态分布 。

Objective The complementarity determining region 3 (CDR3) of T cell receptor (TCR) is the place through which T cells connect to the antigen The lengths and DNA sequences of CDR3s are different according to different T cell clones This leads to a diverse TCR CDR3 repertoires which can reflects the functional status of T cells precisely This study aimed at elucidating the abnormality of TCR β chain variable region (BV) CDR3 repertoires of children with acute B lymphoblastic leukemia, the pathogenesis of leukemia associated with T cell dysfunction and the immuno reconstruction of T cells after the chemotherapy Methods Twelve children aged from 3 to 13 years (average 4 50±3 78 years) with acute B lymphoblastic leukemia before chemotherapy and 8 healthy control donors aged from 6 to 16 years (average 10 30±3 00 years) were enrolled Four of 12 patients were studied for the second time 3 months after complete remission (CR) Reverse transcription polymerase chain reaction (RT PCR) and polyacrylamide sequencing gel electrophoresis were used to detect the diversity of TCR BV CDR3 repertoires of these children Results (1)The expression of BV2 and BV3 in 12 children increased and BV17 and BV18 decreased before the chemotherapy as compared with controls ( P <0 05) There were 4 children with a lower level expression of BV21 before the chemotherapy, and much lower level expression was found after the remission (2) Normally each lane contained eight to ten bands, which represented unique CDR3 sizes for a given TCR BV family Bands differed in size by 3 nucleotides and generally form a Gaussian distribution There were 14% TCR BV families with abnormal CDR3 length distribution in 12 patients before therapy, which was significantly higher than that of controls (5 5%, P <0 05) Restricted CDR3 length distribution was observed in BV14, BV1, BV16, BV20, BV13 1, BV13 2 and BV6 Every abnormal BV family recovered to normal Gaussian distribution 3 months after CR Conclusion T lymphocytes in children with acute B lymphoblastic leukemia revealed markedly skewed repertoires, which suggests the abnormality of T cell functions Most abnormal T cell repertoires recovered to normal Gaussian distribution 3 months after the first CR, which suggests the immunological reconstitution of T cell repertoires