Increased reactive oxygen species and exhaustion of quiescent CD34-positive bone marrow cells may contribute to poor graft function after allotransplants

Abstract

Poor graft function (PGF) is a fatal complication following allogeneic haematopoietic stem cell transplantation. However, the underlying mechanism is unclear. Effective cross-talk between haematopoietic stem cells (HSCs) and bone marrow microenvironment is important for normal haematopoiesis. Normal HSCs reside in a hypoxic bone marrow microenvironment that protects them from oxidative stress that would otherwise inhibit their self-renewal and results in bone marrow failure. Whether an increased level of reactive oxygen species (ROS) causes PGF following allotransplant is unclear. Using a prospective case-pair study, we identified increased levels of ROS in CD34⁺ bone marrow cells in subjects with PGF. Elevated ROS levels was associated with an increased frequency of DNA strand breaks, apoptosis, exhaustion of quiescent CD34⁺ cells and defective colony-forming unit plating efficiency, particularly in the CD34⁺CD38^- fraction. Up-regulated intracellular p53, p21, caspase-3 and caspase-9 levels (but not p38) were detected in CD34⁺ cells, particularly in the CD34⁺CD38^- fraction. To further study the potential role of ROS levels in post-transplant haematopoiesis, CD34⁺ bone marrow cells from subjects with good graft function were treated with H₂O₂. This increased ROS levels resulting in defective CD34⁺ cells, an effect partially reversed by N-acetyl-L-cysteine. Moreover, CD34⁺ bone marrow cells from the donors to subjects with poor or good graft function exhibited comparable haematopoietic reconstitution capacities in the xeno-transplanted NOD-Prkdc^scidIL2rg^null mice. Thus, even if the transplanted donors’ bone marrow CD34⁺ cells are functionally normal pre-transplant, ROS-induced apoptosis may contribute to the exhaustion of CD34⁺ bone marrow cells in subjects with PGF following allotransplant.