Reconstitution of Endothelial Progenitor Cells after Allogeneic Bone Marrow Transplantation in Children with Malignancies.
Authors: Rosti Vittorio , Massa Margherita , Bonetti Elisa , Campanelli Rita , Meli Valentina , Castelnovi Claudia , Lisini Daniela , Pagliara Daria , Mina Tommaso , Bernardo Maria E. , Montagna Daniela , Maccario Rita , Locatelli Franco 
Published: April 2020

Abstract
Evidences have been reported that bone marrow (BM)-derived endothelial progenitor cells (EPCs) circulate in peripheral blood (PB) of healthy subjects. These EPCs seem to play an important role in maintaining homeostasis of vessel walls, participating in both neo-angiogenetic processes and re-endothelization of the wall of injured vessels. The aim of this study was to assess the number and origin of circulating EPCs in PB and BM of children who underwent allogeneic BMT for malignancies. Thirty-five patients with acute lymphoid leukemia (n=18), acute myeloid leukemia (n=13), non Hodgkin lymphoma (n=1), myelodysplastic syndrome (n=1), chronic myeloid leukemia (n=1), and rabdomyosarcoma (n=1) were enrolled in this study. We evaluated PB samples at 21 days, and either PB or BM samples at 45, 60, 90, 120, 180, and 365 days after transplantation. The number of EPCs was evaluated as CD34+VEGFR-2+ or CD34+CD133+VEGFR-2+ cells by cytofluorimetric analysis, and by in vitro culture. PB (n=10) and BM (n=10) samples from age-matched BM donors were analyzed as controls. Donor or recipient origin of EPCs was assessed, by micro-satellite analysis, on at least 10 individually-picked endothelial colonies. The percentage of circulating CD34+VEGFR-2+ cells was significantly lower (p=0.02) in patients tested 21 days after transplant (median 0.01%, 0–1.0) than in PB from controls (median 0.06%, 0–1.3). At the same time point, the percentage of CD34+ co-expressing the CD133 and VEGFR-2 antigens, representing a restricted subset of immature EPCs, was lower, although still not-statistically significant, in patients (median 0.0%, 0.0–4.6) than in controls (median 0.6%, 0.0–15.1); the number of EPC-derived colonies was also significantly lower (p&lt;0.002) in patients at 21 days from BMT (median 4/106 MNC, 0–15) than in controls (median 22/106 MNC, 11–43). Neither the percentage of circulating cell subsets, nor the number of EPC-derived colonies, showed significant modifications during 1-year follow up (ANOVA test). Similar results were obtained when BM samples were analyzed. The percentage of CD34+VEGFR-2+ and CD34+CD133+VEGFR-2+ EPCs was lower in patients tested at 45 days after transplantation than in controls (p=0.03 and p&lt;0.04, respectively). The number of EPC-derived colonies was also lower (p= 0.001) than that found in BM from controls. At subsequent time points, no significant variation of these parameters was found (ANOVA test). Conditioning regimen, or occurrence of GvHD did not influence the percentage of EPCs in either PB or BM. Microsatellite analysis was performed on EPC-derived colonies of 4 patients, at time points ranging from 45 days to 9 months after the allograft. All the analyzed colonies were of donor origin. In conclusion, circulating and BM EPCs are detectable in children given BMT from 21 days up to 1 year after transplant. These cells are derived from donor BM indicating that EPCs are transferred with the graft into the recipient. More cases and longer follow up studies are needed to assess whether these cells can be correlated with clinical outcome of the transplanted patients.c