Li Yangqiu, Chen Shaohua, Yang Lijian, Li Bo, Chan John Yeuk-Hon, Cai Dongqing
Institute of Hematology, Medical College, Jinan University, Guangzhou 510632, China.
Transpl Immunol. 2009 Jan;20(3):155-62. doi: 10.1016/j.trim.2008.10.010. Epub 2008 Nov 12.
Umbilical cord blood (CB) has been used as a valuable source of hematopoietic stem cells for allogeneic transplantation, specific CTL response and immunotherapy for decades. We previously analyzed the distribution and clonality of T-cell receptor alpha and beta variable region (TRAV) and (TRBV) of the subfamily T cell receptors in T cells from umbilical cord blood. Recent data indicated that gammadelta(+) T cells may play an important role in mediating the graft versus leukemia effect after stem cells transplantation and in anti-cancer response. In order to further characterize the repertoire of CB T-cells, the frequency of alphabeta(+) and gammadelta(+) T cells were examined in CB by FACS. The CDR3 size of 4 TRGV and 8 TRDV subfamily genes were analyzed in mononuclear cells (MCs) from 16 CB samples, using RT-PCR and genescan technique. To determine the expression level of TRGV subfamily genes, we performed quantitative analysis of TRGVI-III subfamilies by real-time PCR. Low percentage of CD3(+)TCRgammadelta(+) cells was observed in CB. The frequency of expression in TRGVI, TRGVII and TRGVIII in CBMCs was 93.75%, 81.25% and 56.25%, respectively. The mean value of the number of expressed TRDV subfamilies in CBMCs is higher than that from adult peripheral blood (PB) group. The frequently expressed members in CB were TRDV1 (100%), TRDV2 (93.75%), TRDV8 (93.75%) and TRDV3 (81.25%), respectively. The frequencies of TRDV5 and TRDV8 in CBMCs were significantly higher than those from PBMCs. Most of the PCR products of TRGV and TRDV subfamilies from 10 CB samples displayed polyclonal rearrangement pattern, whereas one or two PCR products from 6 CB samples showed oligoclonality or biclonality. In contrast, PCR products from 9 of 10 adult healthy controls contained at least an oligoclonal peak in different TRGV or TRDV subfamilies respectively. The pattern of TRGV subfamily expression level in CBMCs was TRGVI>TRGVIII>TRGVII, and in contrast, TRGVII>TRGVI>TRGVIII was found in PBMCs. In conclusion, our results indicate polyclonal and more diverse TRDV segment usage in CB gammadelta(+) T-cells. The pattern of TRGV expression levels in CB T cells was found to be quite different from the one in PB T cells. These findings are apparently the first report regarding the repression pattern of TRGV repertoire in CB. It also provides a detailed profile of the global TRGV and TRDV repertoire and TRGVI-III expression levels in cord blood T cells in Chinese subjects. The biological significance of the differences observed between CB and PB is at present obscure. However, this study will definitively contribute to understand the cellular immune features better and to exploit more efficiently the therapeutic potentials of CB.
几十年来，脐带血一直是用于异基因移植、特异性CTL反应和免疫治疗的造血干细胞的宝贵来源。我们之前分析了脐带血T细胞中T细胞受体α和β可变区（TRAV）和（TRBV）亚家族T细胞受体的分布和克隆性。最近的数据表明，γδ(+) T细胞可能在干细胞移植后的移植物抗白血病效应和抗癌反应中发挥重要作用。为了进一步表征脐带血T细胞的库，通过流式细胞术检测了脐带血中αβ(+)和γδ(+) T细胞的频率。使用RT-PCR和基因扫描技术分析了16份脐带血样本的单核细胞（MCs）中4个TRGV和8个TRDV亚家族基因的CDR3大小。为了确定TRGV亚家族基因的表达水平，我们通过实时PCR对TRGVI-III亚家族进行了定量分析。在脐带血中观察到CD3(+)TCRγδ(+)细胞的百分比很低。脐带血单个核细胞（CBMCs）中TRGVI、TRGVII和TRGVIII的表达频率分别为93.75%、81.25%和56.25%。脐带血单个核细胞中表达的TRDV亚家族数量的平均值高于成人外周血（PB）组。脐带血中频繁表达的成员分别为TRDV1（100%）、TRDV2（93.75%）、TRDV8（93.75%）和TRDV3（81.25%）。脐带血单个核细胞中TRDV5和TRDV8的频率显著高于外周血单个核细胞（PBMCs）。10份脐带血样本中TRGV和TRDV亚家族的大多数PCR产物显示多克隆重排模式，而6份脐带血样本中的一两个PCR产物显示寡克隆性或双克隆性。相比之下，10名成人健康对照中的9份PCR产物分别在不同的TRGV或TRDV亚家族中至少包含一个寡克隆峰。脐带血单个核细胞中TRGV亚家族表达水平的模式为TRGVI>TRGVIII>TRGVII，相反，在外周血单个核细胞中发现TRGVII>TRGVI>TRGVIII。总之，我们的结果表明脐带血γδ(+) T细胞中TRDV片段的使用是多克隆的且更加多样化。发现脐带血T细胞中TRGV表达水平的模式与外周血T细胞中的模式有很大不同。这些发现显然是关于脐带血中TRGV库抑制模式的首次报告。它还提供了中国受试者脐带血T细胞中全球TRGV和TRDV库以及TRGVI-III表达水平的详细概况。目前，脐带血和外周血之间观察到的差异的生物学意义尚不清楚。然而，这项研究将无疑有助于更好地理解细胞免疫特征，并更有效地开发脐带血的治疗潜力。
