The Impact of Rhg-CSF on the LFA-1 Conformation of CD4+T Cells in Mobilized Hematopoietic Stem Cell Allografts

In hemotopoietic stem cell transplantation (HSCT), the primary effects of G-CSF on cells of the hemotopoietic system include stimulation of proliferation and differentiation of HSCs, acceleration of neutrophil reconstitution after HSCT and mobilization of bone marrow HSCs into the peripheral blood. In recent years, several investigators have unraveled that G-CSF-mediated immune regulation including the switching T cell cytokine secretion profile to Th2 response, the altering adhesion activity of CD4+T cells to ICAM-1 and so on. Most of these cellular interactions of T cells are dependent on the integrin leukocyte function associated antigen-1 (LFA-1, α_Lβ_2, CD11a/CD18) binding to intercellular adhesion molecule-1 (ICAM-1). Integrin α_Lβ₂ adopts a bent, compact conformation in their low affinity state and an extended conformation in the high affinity state. The epitope of MEM148 is only expressed on the free β₂ integrin subunit but is masked on LFA-1. The monoclonal antibody MEM148 reacts with an epitope exposed on free human CD18 chain as well as on high affinity state of LFA-1 that represents a reporter for α_Lβ₂ active conformation. Our previous studies have demonstrated that rhG-CSF mobilization decreased the adhesion activity of CD4+T cells to ICAM-1, but did not find that rhG-CSF mobilization had effect on the level of CD11a expression. rhG-CSF maybe has effect on conformational change of LFA-1 on CD4+T cells. In order to explore the impact of rhG-CSF mobilization on LFA-1 conformation of CD4+ T cells, human CD4+T cells were isolated from peripheral blood mononuclear cells by positive selection with Miltenyi MACS. Isolated CD4+T cells were activated by OKT3+ICAM-1 and PMA+Ion separately. The slides with CD4+T cells were incubated with FITC-CD11a and FITC-MEM148 mAb, then were examined by using fluorescence microscope. The results show that the level of CD25, CD69 expression and MEM148 epitope exposure on activated CD4+T cells were significantly higher compared with unactivated CD4+T cells. rhG-CSF mobilization had no effect on the expression of CD11a, but could decrease expression of CD25, CD69 and exposure of MEM148 epitope on activated CD4⁺T cells. The percentage of LFA-1 polarized CD4+T cells before and after rhG-CSF mobilization was 85.32%, 61.86% respectively (p <0.01). No MEM148 epitope exposure on unactivated CD4+T cells was detected, but MEM148 epitope exposure on activated CD4+T cells before and after rhGCSF mobilization was 63.63%, 32.79% respectively (p <0.01). Overall, these data suggest that rhG-CSF can affect on the activation of CD4+T cells in mobilized hematopoietic stem cell allografts by altering the conformation of LFA-1.