Functional and Phenotypic Characterizations of Granulocyte-Colony Stimulating Factors Mobilzed CD34+ Cells.
Authors: Bijou Fontanet , Hammoud Muhamed , Vlaski Marija , Duchez Pascale , Milpied Noel-Jean , Lafarge Xavier , Chevaleyre Jean , Ridha Bouzgarrou , Dazey Bernard , Boiron Jean-Michel , Ivanovic Zoran 
Published: October 2019

Abstract
Abstract 2143
Poster Board II-120

Introduction:
Since the mid 1985 the combination of hematopoietic growth factors and/or chemotherapy is applied to allow mobilization of CD34+ cells from marrow to the blood. (pHSC). Once harvested, this cell population, containing both progenitors (pHSC) and stem cells (HSC), has become the common source for autologous or/and allogeneic transplantations procedures. Indeed, the new development of cell-based therapies in hematologic settings requires the understanding of mechanism involved in HSC migration and cell cycle kinetics during mobilisation.

Materials and Methods:
With respect to the percentage and absolute number of CD34+ cells after Granulocyte-Colony Stimulating Factor mobilisation (G-CSF) the patients (Pts) and healthy donors were divided in two groups: Good mobilizers (GM) and poor mobilizers (PM). We studied in their CD34+ cells: cell cycle (Ki67/IP staining), clonogenic and ex vivo expansion potential, aldehyde deshydrogenase activity (Aldefluor), expression of some adhesion molecules relevant for CD34+cells mobilisation as: VLA-4, LFA-1 and CXCR4.

Results:
Twenty Two Pts (Myeloma=13 and Lymphoma=9) and 9 healthy donors underwent G-CSF mobilisation with or without chemotherapy. CD34+ cells were purified using the MACS cell isolation kit and Mini-Macs columns obtaining a purity of more than 88%. Based on the percentage of circulating CD34+, 2 populations of subjects were identified (<0.1 %: PM and >0.1%: GM). Total white blood cells number was higher in PM arm (48.8 G/l) than in GM arm (36.75) (p= 0.03). The median pCD34+ cell was higher in GM (60: 31-153) than in PM (35: 11-64) (p= 0.001). Interestingly, PM showed more G0 cells than GM with mean percentages 46±10 and 31±10 respectively (p=0.004). Indeed, the percentage of the S/G2/M fraction of CD34+ cells was higher in GM (2±0.9) than in PM (0.69±0.7) (p= 0.002). Moreover, there was a trend to observe a higher total nucleated cells fold expansion potential in PM arm 27±11 versus 17±13 in GM (p=0.06). It should be stressed that those results were observed also when Pts and healthy donors were studied separately. Analysis of ALDH activity revealed an unexpected population of CD34+ cells that were ALDH+ despite their CD133 negativity. These CD34+ALDH+CD133− cells were present in both patients and healthy donors but were more pronounced in pts (38% in PM versus 9% in GM; p= 0.01). Moreover, GM among healthy donors showed more cells expressing VLA-4 (87 % versus 61 % in PM arm, p= 0.05)

Conclusion
These results suggest important phenotypic and functional variations in CD34+ cells between PM and GM (cell cycle, expression of CD133 by ALDH+ cells) as well as some differences between the healthy donors and pts (VLA-4 expression and clonogenic capacity).

Disclosures:
No relevant conflicts of interest to declare.