The Frequency and Clonality of γδ+T Cells, distribution of TRGV and TRDV Repertoire in Cord Blood T Cells

Our previous study has showed the feature of distribution and clonality of T-cell receptor alpha variable region (TRAV) and TRBV subfamily T cells. Recently, data indicated the γδ+T cells may play an important role in mediated graft versus leukemia effect after stem cells transplantation and anti-cancer response. In order to farther characterize the repertoire of CB T-cells, the frequency of αβ+ and γδ+T cells were detected in cord blood (CB) by FACS, the CDR3 size of 4 TRGV and 8 TRDV subfamily genes were analyzed in mononuclear cells (MCs) from 16 cord blood samples, using RT-PCR and genescan technique. Ten healthy individuals served as control. To determine the expression level of TRGV subfamily genes, quantitative analysis of TRGV I–III subfamilies was performed by real-time PCR. Low percentage of CD3⁺TCRγδ⁺ cells (1.33±0.58%) was observed in CB. The frequency expression was found in TRGVI (93.75%) and TRGVII (81.25%) in CBMCs, whereas the TRGVIII was detected in 9 out of 16 samples (56.25%). In TRDV subfamilies, the mean value of the number of expressed subfamilies in CBMCs is higher than that from adult peripheral blood (PB) group. The most frequently expressed in CB were TRDV1 (100%), TRDV2(93.75), TRDV8 (93.75%) and TRDV3 (81.25%). The frequency of TRDV5 and TRDV8 in CBMCs were significant higher that that from PBMC. The most PCR products of TRGV and TRDV subfamilies from 10 cord blood displayed polyclonal rearrangement pattern, whereas one or two PCR products from 6 CB samples displayed 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. In the present study, all PCR products form TRGVIV were showed monoclonality, the direct sequencing results showed that the V region of TRGVIV joint direct to C region, indicating that there is no rearrangement in TRGVIV and it is a pseudogene. The sequence data have been submitted to genebank and the Accession Number is EU395807. The pattern of TRGV subfamily expression level in CBMCs was TRGVI>TRGVIII>TRGVII, in contrast, TRGVII>TRGVI>TRGVII was found in PBMCs. In conclusion, our results indicate polyclonal and more diverse TRDV segment usage in CB γδ⁺ T-cells. The pattern of TRGV expression levels was different between T cells from PB and CB.