MACROPHAGE DEPLETION PROMOTES XENOGENEIC HEMATOPOIETIC CHIMERISM

Abstract

1006 Background: The successful establishment of xenogeneic tolerance through hematopoietic stem cell engraftment is restricted by the rapid disappearance of these cells in the recipient following infusion. Therefore, the goal of these studies was to develop methods to increase survival of pig hematopoietic cell in a xenogeneic recipient. We developed and tested the hypothesis that the mononuclear phagocyte system was responsible for the rapid clearance of infused pig hematopoietic cells. Methods: Balb/c or SCID mice were conditioned with a dose of 3Gy whole body irradiation (WBI), heparin, and cobra venom factor (CVF) on day 0. On various days, mice were injected with medronate-encapsulated liposomes or control blank liposomes, followed by i.v. infusion of miniature swine bone marrow cells. As a means of measuring mononuclear phagocyte function, mice were injected i.v. with FITC-labeled latex beads. The kinetics of bead clearance from the circulation were measured by flow cytometry. To study cell trafficking, swine or autologous bone marrow cells were loaded with a vital fluorochrome. Bone marrow cells (5×107) were then infused i.v. The kinetics of the disappearance of infused cells from the circulation and the trafficking through the spleen and into the bone marrow were determined by flow cytometry. Hematopoietic cells were recovered from the bone marrow at various times and CFU and LTC-IC analyses were performed. Results: Medronate-encapsulated liposomes are non-toxic in mice at levels that deplete mononuclear phagocytes. Depletion of mononuclear phagocytes in Balb/c mice as well as in SCID mice increases the accumulation pig hematopoietic cells in the bone marrow by 10-fold when measured 24hr post bone marrow cell infusion. Following phagocyte depletion, the accumulation of pig cells in the bone marrow of Balb/c mice is approximately two-fold less than that observed following autologous bone marrow cell infusion. LTC-IC assays indicate an increased presence of pig progenitors in the bone marrow of mice which were treated pre- and post- bone marrow infusion with medronate-liposomes. Experiments to investigate the effect of phagocyte depletion on pig cell chimerism in primates are in progress. Conclusion: We have found that mononuclear phagocytes play a major role in the elimination of pig hematopoietic cells in mice. Depletion of mononuclear phagocytes in mice leads to a significant increase in pig hematopoietic chimerism and this liposome treatment is not toxic to pig hematopoietic progenitors. This work was supported by BioTransplant, Inc.