Bone Marrow Transplantation (2002) 29, 557–562  2002 Nature Publishing Group All rights reserved 0268–3369/02 $25.00 www.nature.com/bmt CD34  cells Persistent decrease in proliferative potential of marrow CD34  cells exposed to early-acting growth factors after autologous bone marrow transplantation J Domenech 1 , G Cartron 1,2 , N Clement 1 , M-H Estienne 1 , O Herault 1 , D Truglio 1 , L Benboubker 2 , F Roingeard 1 , I Desbois 3 , P Colombat 2 and C Binet 1 1 UPRES-EA 3249, Laboratory of Hematology, University Hospital of Tours, Tours, France; 2 UPRES-EA 3249, Department of Medical Oncology and Blood Diseases, University Hospital of Tours, Tours, France; and 3 UPRES-EA 3249, Regional Blood Bank, University Hospital of Tours, Tours, France Summary: Post-graft hematopoiesis is characterized by long-term quantitative deficiency in marrow progenitor cells in both autologous and allogenic settings. In order to evaluate the function of post-graft progenitor cells, the proliferative capacity of marrow CD34 + cells was evalu- ated in 10 patients 6 months after autologous bone mar- row transplantation (ABMT) for non-Hodgkin’s lym- phoma and compared to that of 10 patients before ABMT and 10 normal controls. Immuno-selected CD34 + cells were cultured for 7 days in liquid serum-free medium with a combination of early-acting GF con- sisting of stem cell factor, IL-3 and IL-1. Clonogenic efficiency of unselected cells for CFU-GM and BFU-E was decreased in post-graft patients compared to pre- graft and control patients. However, clonogenic efficiency of selected CD34 + cells for CFU-GM was not different in post-graft, pre-graft and control patients but BFU-E values of post-graft patients remained lower than those of control patients. Decreased percentages of CD34 + CD38 - cells were observed in both post-graft and pre-graft patients while those of CD34 + c-kit + cells were similar in all three patient groups. After 7-day liquid culture, expansion yields of total progenitor cells were significantly lower in post-graft patients (147 ± 28%) than in pre-graft (255 ± 27%) and control patients (246 ± 23%). Post-graft deficiency in progenitor cell expansion was particularly marked for BFU-E (61 ± 24%) compared to pre-graft patients (220 ± 82%) and to controls (349 ± 82%). These results indicate impaired proliferative potential of marrow CD34 + cells several months after ABMT involving erythroid pro- genitor cells and/or commitment towards erythroid lin- eage from a more immature stage (pre-CFU). Bone Marrow Transplantation (2002) 29, 557–562. DOI: 10.1038/sj/bmt/1703512 Correspondence: Dr J Domenech, Laboratoire d’He ´matologie, Ho ˆpital Bretonneau, 2, Bd Tonnelle ´, 37044 Tours Cedex, France Received 8 October 2001; accepted 4 January 2002 Keywords: autologous bone marrow transplantation; post-graft hematopoiesis; CD34 + progenitor cells; prolifer- ative capacity; early-acting growth factors Over the past 15 years, high-dose therapy has been widely used in onco-hematology to improve the efficacy of anti- tumor treatments. The high myelotoxicity of such regimens requires hematopoietic rescue by transplantation of allo- geneic, syngeneic or autologous progenitor and stem cells. To ensure full and stable hematopoietic reconstitution, these grafts must contain sufficient amounts of precursor and progenitor cells responsible for early engraftment and primitive stem cells for late engraftment. 1,2 Normalization of peripheral blood cell (PBC) counts is usually considered to estimate successful hematopoietic engraftment but this criterion does not really reflect full hematopoietic reconsti- tution. Indeed, as shown after allogeneic bone marrow transplantation (BMT), 3 we have reported a decrease in marrow hematopoietic progenitor cell (HPC) frequency for up to 4 years after autologous BMT (ABMT) 4 whereas most of the patients studied normalized PBC counts within 4–6 weeks. A long-lasting impairment of the hematopoietic system could lead to the emergence of secondary hemo- pathies such as myelodysplastic syndrome or acute leuke- mia whose actuarial risk at 10 years has been estimated to be above 10% in several reports. 5–8 Although we noted that post-graft deficiency of HPC from the mononuclear cell (MNC) fraction mainly involved the erythroid lineage, 4 it could not be attributed to an abnormal response of erythroid progenitor cells to stem cell factor (SCF) that increased their clonogenicity as well as erythroid colony size. 9 How- ever, using a Dexter-type long-term marrow culture (LTMC) system, 10 we found a decrease in CFU-GM pro- duction for up to 1 year after ABMT. 11,12 This correlated with low initial counts of erythroid progenitor cells and suggested abnormal proliferative capacity at the primitive cell level. 11 The aim of this study was to evaluate the proliferative capacity of immature HPC after ABMT in response to early-acting growth factors (GF) using a short-term stroma-