Stem cell transplantation eliminates alloantibody in a highly sensitized patient.

Highly sensitized patients are forced to stay on transplant waiting lists for many years and ultimately may never find a donor. Peripheral blood stem cell (PBSC) transplantation may provide a strategy to decrease host alloreactivity through the production of a chimeric state. We investigated alloreactivity and chimerism in a highly sensitized 40-year-old patient with sickle cell disease who underwent a nonradiation based conditioning regimen consisting of fludarabine, ATG, and high dose melphalan, for allogeneic stem cell transplant. Host monocytes and lymphocytes became donor in origin by day 14. PRA, initially 100% pretransplant, fell to 0 by day 263. Anti-red blood cells antibody became undetectable by day 152. The use of a new nonradiation-based conditioning regimen enabled successful engraftment of allogeneic donor PBSCs and the elimination of alloantibody. As new less toxic conditioning regimens are developed, PBSC transplantation might provide a new solution to allosensitization.

Delayed effects of rhG-CSF mobilization treatment and apheresis on circulating CD34+ and CD34+ Thy-1dim CD38- progenitor cells, and lymphoid subsets in normal stem cell donors for allogeneic transplantation.

Allogeneic transplantation of peripheral blood progenitor cells (PBPC) is emerging as a new stem cell transplant modality. Rather than undergoing general anesthesia for bone marrow harvest, normal blood stem cell donors are subjected to rhG-CSF mobilization treatment followed by single or multiple apheresis. Whereas the effects of cytokine treatment and apheresis on stem cell peripheralization and collection have been described, little is known about delayed effects of rhG-CSF treatment and apheresis on a normal hematopoietic system, and there are no long-term data that address safety issues. Ten normal, patient-related donors underwent a 3 or 4 day rhG-CSF (filgrastim) treatment (12 micrograms/kg/day) followed by single or tandem apheresis. We monitored peripheral blood (PB) cellularity including CD34+ and lymphoid subsets at baseline, during cytokine treatment, prior to apheresis, and at days 2, 4, 7, 30 and 100 post-apheresis. The PB progenitor cell concentration peak prior to apheresis was followed by a nadir by day 7 and normalized by day 30, with the exception of the most primitive CD34+ Thy-1dim CD38- progenitor subset that reached a nadir by day 30. Lymphoid subsets such as CD3, 4, 8, suppressor cells (CD3+ 4- 8- TCR+ alpha beta), and B cells (CD19+) showed a similar pattern with a nadir concentration by day 7, followed, except for B cells, by a rebound by day 30 and subnormal counts at day 100. The PB concentrations of hemoglobin and platelets dropped mainly due to the apheresis procedure itself, and normalized by day 30. With cytokine treatment, the PB alkaline phosphatase and lactate dehydrogenase concentrations increased 2.2- and 2.8-fold, respectively, over baseline, and returned to normal range by day 30. Based on the preliminary nature of this study, the clinical relevance of these findings is still unclear.

Simultaneous use of rhodamine 123, phycoerythrin, Texas red, and allophycocyanin for the isolation of human hematopoietic progenitor cells.

The supravital, mitochondrial specific dye Rhodamine 123 (R123) was used in conjunction with three monoclonal antibodies to isolate a population of human bone marrow (BM) cells enriched for hematopoietic progenitor cells. BM cells stained with phycoerythrin-HLA-DR, Texas red-CD34, allophycocyanin-CD15, and R123 were fractionated using four-color immunofluorescence cell sorting. Cells expressing CD34 but not HLA-DR and CD15 (CD34+ HLA-DR- CD15-) were subdivided according to their reactivity with R123 into quiescent, R123 dull (R+) or cycling, R123 bright (R++) subpopulations. Morphological analysis and hematopoietic progenitor cell assays indicated that CD34+ HLA-DR- CD15- R+ cells contained larger numbers of blast cells and colony forming units than CD34+ HLA-DR- CD15- R++ cells. The flow cytometer settings used to accommodate the detection of the R123 fluorescence in combination with that of three other fluorochromes are described.