Granulocyte colony-stimulating factor affects the distribution and clonality of TRGV and TRDV repertoire of T cells and graft-versus-host disease

Abstract

Background: The immune modulatory effect of granulocyte colony-stimulating factor (G-CSF) on T cells resulted in an unexpected low incidence of graft-versus-host disease (GVHD) in allogeneic peripheral blood stem cell transplantation (allo-PBSCT). Recent data indicated that gamma delta+ T cells might participate in mediating graft-versus-host disease (GVHD) and graft-versus-leukemia (GVL) effect after allogeneic hematopoietic stem cell transplantation. However, whether G-CSF could influence the T cell receptors (TCR) of gamma delta+ T cells (TRGV and TRDV repertoire) remains unclear. To further characterize this feature, we compared the distribution and clonality of TRGV and TRDV repertoire of T cells before and after G-CSF mobilization and investigated the association between the changes of TCR repertoire and GVHD in patients undergoing G-CSF mobilized allo-PBSCT.

Methods: The complementarity-determining region 3 (CDR3) sizes of three TRGV and eight TRDV subfamily genes were analyzed in peripheral blood mononuclear cells (PBMCs) from 20 donors before and after G-CSF mobilization, using RT-PCR and genescan technique. To determine the expression levels of TRGV subfamily genes, we performed quantitative analysis of TRGVI~III subfamilies by real-time PCR.

Results: The expression levels of three TRGV subfamilies were significantly decreased after G-CSF mobilization (P = 0.015, 0.009 and 0.006, respectively). The pattern of TRGV subfamily expression levels was TRGVII >TRGV I >TRGV III before mobilization, and changed to TRGV I >TRGV II >TRGV III after G-CSF mobilization. The expression frequencies of TRGV and TRDV subfamilies changed at different levels after G-CSF mobilization. Most TRGV and TRDV subfamilies revealed polyclonality from pre-G-CSF-mobilized and G-CSF-mobilized samples. Oligoclonality was detected in TRGV and TRDV subfamilies in 3 donors before mobilization and in another 4 donors after G-CSF mobilization, distributed in TRGVII, TRDV1, TRDV3 and TRDV6, respectively. Significant positive association was observed between the invariable clonality of TRDV1 gene repertoire after G-CSF mobilization and low incidence of GVHD in recipients (P = 0.015, OR = 0.047).

Conclusions: G-CSF mobilization not only influences the distribution and expression levels of TRGV and TRDV repertoire, but also changes the clonality of gamma delta+ T cells. This alteration of TRGV and TRDV repertoire might play a role in mediating GVHD in G-CSF mobilized allo-PBSCT.

Figure 1

The expression frequencies of TRGV and TRDV subfamilies in PBMCs from 20 donors before and after G-CSF mobilization.

Figure 2

The pattern of TRGV I-III expression levels in PBMCs from 20 donors before and after G-CSF mobilization.

Figure 3

Distribution and clonality of TRGV and TRDV subfamilies in PBMCs from 20 donors (D1-D20). (A) before G-CSF mobilization; (B) after G-CSF mobilization. GVHD*: the status of graft-versus-host disease of the corresponding recipients after allogeneic peripheral blood stem cell transplantation; a: acute graft-versus-host disease; c: chronic graft-versus-host disease.

Figure 4

Changes of clonality of TRGV and TRDV subfamilies in PBMCs from 7 donors (D5, D7, D8, D3, D4, D10 and D11) before and after G-CSF mobilization. (A) Oligoclonal expansion changed to polyclonality or negative after mobilization; (B) Polyclonal or negative expansion changed to oligoclonality after mobilization.