Mechanisms underlying the human cytomegalovirus infection of human hematopoietic cells

The aim of this study was to investigate the infection of human hematopoietic stem cells (HSC) with human cytomegalovirus (HCMV). Until recently, all studies that investigated the infection of human hematopoietic cells have of necessity been performed in vitro for obvious ethical reasons. We used transplantation into the preimmune human/sheep xenograft model to evaluate the mechanisms underlying the role of HSC in the mediation of HCMV infection in man. CD34 +Lin− HSC were isolated by fluorescence-activated cell sorting (FACS) from the bone marrow (BM) of normal human HCMV positive and negative donors, and the latter was subjected to HCMV infection. Viral HCMV DNA was detected in both populations by nested-polymerase chain reaction (nested-PCR) and fluorescent in situ hybridization (FISH) prior to transplant. Although human hematopoietic cell engraftment was detected in BM and peripheral blood (PB) at 5 and 7 months post transplant with levels of engraftment varying from 0.13–22%, all sheep were negative for HCMV. To ensure that our inability to detect HCMV-positive cells was not due to HCMV becoming inactivated within the sheep, experiments were performed in which uncharacterized HCMV-positive BM stroma was transplanted into the sheep. The heart, spleen, kidney, thymus and skin of transplanted sheep were found to be positive for both HLA-DQα and HCMV. These results suggest that the long-term repopulating HSC cannot be infected with HCMV and raises the possibility that human blood cells become infected at the level of the committed hematopoietic progenitor. This was supported by the study of the infection of human megakaryocytes with HCMV. Megakaryocytic cells and their early precursors were shown to be susceptible to in vitro infection with HCMV. Magnetic column-purified CD42+ megakaryocytes were positive for HCMV by performing two-color fluorescent staining with antibodies directed against CD42 and HCMV pp65 or I/E antigen. These findings were confirmed by infecting megakaryocytes with a laboratory strain of HCMV containing the β-galactosidase (β-gal) gene. Production of infectious virus was observed in cultures of infected megakaryocytes suggesting that HCMV can complete its life cycle. Our results demonstrate that the short-term repopulating stem cells may be the target of HCMV infection.