ALBERT

Description: To study the complex pathophysiology of aGvHD in allogeneic hematopoietic cell transplantation (HCT) we transplanted transgenic luciferase expressing T cell populations into lethally irradiated HCT recipients (murine MHC major mismatch model, H-2q into H-2d). Tracking of light emitting donor T cells in living animals and detailed studies by multi color immunofluorescence microscopy (IFM) and FACS revealed the tight links of spatial and temporal evolution in this complex immune process. Donor derived T cells migrate to T cell areas in lymphoid tissues within a period of 12 hours. In the initial periods donor CD4+ T cells appear first with CD8+ T cell infiltration at later time points. Donor T cells start proliferating in lymphatic tissues on day 2 after transfer, as observed by BrdU stainings. Although alloreactive T cells are similarly activated in all lymphoid organs, they only up-regulate gut homing molecules after more than 5 cell divisions (CFSE proliferation analysis by FACS) in certain lymphoid organs (Peyer’s patches, mesenteric LN and spleen). Abruptly on day 4 after HCT, T cells migrate into intestinal sites. These findings strongly suggested, that specific priming sites are required for alloreactive T cells to induce a distinct type of tissue tropism in GvHD. In contrast to previous reports peformed without host conditioning, depletion of certain lymphoid organs (e.g. Peyer’s patches) before HCT or antibody blocking experiments did not control aGVHD. BLI showed, that anti-L-selectin or anti-MAdCAM-1 antibody treatment alone or in combination was effective in blocking donor T cell migration to lymph nodes and Peyer’s patches, while redirecting these cells to liver and spleen. Subsequently cells proliferated predominantly in the spleen until day 3 after HCT. Surprisingly we observed a full picture of gut infiltration on day 4 and skin involvement on day 5–6, similar in dynamics and strength to the aGvHD isotype control group. These findings demonstrated, that other lymphoid organs can functionally compensate for inducing gut and skin homing of alloreactive T cells. Of importance, we demonstrated that T cells that lacked homing molecules for secondary lymphoid organs had alloreactive properties in vitro, yet did not cause aGVHD in vivo. In summary, the activation of alloreactive T cells in specific sites throughout the body is complex and involves the acquisition of homing molecule expression. Transplantation of T cells with defined homing properties therefore, appears to be a promising alternative in conferring protective immunity early after HCT without the risk of aGvHD.

Description: DiGeorge syndrome (DGS) is a congenital disorder characterized by facial dysmorphies, hypoparathyroidism, cardio-vascular malformations and variable T-cell deficiency due to thymic hypoplasia. We present a male infant with complete absence of T-cells and thymus successfully treated by repeat blood lymphocyte transfusions. According to previous experience of bone marrow transplantation in a similar patient, T-cell reconstitution is possible by peripheral expansion of mature donor cells, independent of stem cell engraftment. Our patient received at the age of four months an initial transfusion of blood lymphocytes from his HLA-identical brother, containing 1x108 T-cells/ kg. Ten days later he developed mild cutaneous GvHD, which resolved spontaneously without immunosuppressive therapy. Lymphocyte transfusions were repeated three times at 4 weeks intervals without reappearance of GvHD. T-cells developed within two months after the first transfusion and since then remain at levels between 500–1000/μl with a proportion of 25% disclosing a CD45RA+ phenotype. In chimerism analysis T-cells are exclusively of donor origin. The T-cell receptor repertoire is similarly diverse as in the donor. After more than 2 years of follow up the boy is not suffering from severe infections and shows no signs of autoimmune disease. Following immunization, antibody titres normalized. Conclusion: In infants with complete DGS transfusions of blood lymphocytes from a matched family donor may lead to long-lasting engraftment of functional T cells and stable chimerism and represents a potentially helpful therapeutic option.

Description: Several transcription factors have been implicated as playing a role in myelopoiesis. PU.1, an ets-family transcription factor, is required for the development of myeloid and lymphoid lineages, whereas the transcription factor CCAAT–enhancer binding protein family member C/EBPα is essential for granulocyte development. We present here the first evidence that C/EBPα blocks the function of PU.1. PU.1 and C/EBPα interact physically and colocalize in myeloid cells. As a consequence of this interaction, C/EBPα can inhibit the function of PU.1 to activate a minimal promoter containing only PU.1 DNA-binding sites. We further demonstrate that the leucine zipper in the DNA-binding domain of C/EBPα interacts with the β3/β4 region in the DNA-binding domain of PU.1 and as a result displaces the PU.1 coactivator c-Jun. Finally, C/EBPα blocks PU.1-induced dendritic cell development from CD34+ human cord blood cells. The functional blocking of PU.1 by C/EBPα could be the mechanism by which C/EBPα inhibits cell fates specified by PU.1 and directs cell development to the granulocyte lineage.