Blood Traffic of Endothelial, Mesenchymal, Hematopoietic and Lin-/CD45-CD34+AC133+CXCR4+ Progenitor Cell Subsets Show Different Patterns in Pre-Fibrotic and Fibrotic Primary Myelofibrosis
Authors: Ojeda-Uribe Mario , Desterke Christophe , Jung Laura , Bories Pierre , Sovalat Hanna , Mauvieux Laurent , Eisenmann Jean Claude , Lioure Bruno , Le Bousse-Kerdiles Marie-Caroline 
Published: November 2019

Abstract
Introduction. Primary myelofibrosis (PMF) can be associated to increased bloodstream traffic of some adult progenitor cells. The extramedullary hematopoiesis developed in this setting might remind some features of the fetal hematopoiesis.
Material and methods. Some evidence of this hypothesis was searched by flow-cytometry and molecular analysis of blood samples from 12 untreated patients with PMF, 6 pre-fibrotic (PF) and 6 fibrotic (F). The PF or F phase was defined according to the WHO and Thiele histopathology scores.
Results. As expected we detected high numbers of CD34+ cells (258x103±402x103/ml; range 1.2-1218x103/ml) as well as small mean numbers of Lin-/CD45-CD34+AC133+CXCR4+ progenitor cells (130±275/ml; range 0-882/ml) that are related to VSELs (very-small embryonic-like stem cells), which have been described as putative pluripotent adult stem cells. When we looked at some embryonic molecular markers by RT-PCR, NANOG and OCT4 expression was detected in the MNC fraction of all the patients tested (hES and CPRE2 cell lines were the positive and negative controls). OCT4 expression was half the level of hES being stronger in F-PMF than in PF-PMF. In all the tested patients NANOG expression was similar to that of hES, whereas SOX2 and LIN28 were not expressed in most patients. Regarding other circulating progenitor cells we observed that PF-PMF had higher mean values of progenitor endothelial cells (PEC) (130±271/ml; range 0-683) than F-PMF (7±16/ml; range 0-37). Mesenchymal progenitor cells (MPC) had also higher mean values in PF-PMF (289±452/ml; range 0-1165) than in F-PMF (111±199/ml; range 0-458). Conversely, Lin-/CD45-CD34+AC133+CXCR4+ cell subset detection was higher in F-PMF (185±381/ml; range 0-882) than in PF-PMF (38±60/ml; range 0-140). Patients V617F-JAK2pos presented lower levels of PEC and MPC (16±19/ml and 43±61/ml respectively) than those V617F-JAK2neg (173±340/ml and 515±470/ml respectively). Similar numbers of Lin-/CD45-CD34+AC133+CXCR4+ cells were observed in both V617F-JAK2pos and V617F-JAK2neg patients (133±331/ml and 123±115/ml respectively).
Patients with elevated IPSS (>3) showed higher levels of PEC, MPC and Lin-/CD45-CD34+AC133+CXCR4+ cells (respectively 180±335/ml, 334±560/ml and 256±422/ml) than those with low IPSS (<2) (respectively 12±16/ml, 134±183/ml and 46±92/ml). Based on these findings we were able to define some putative patterns of bloodstream progenitor cell traffic. Thus, PEC appeared to circulate more in PF-PMF, V617F-JAK2neg with a high IPSS. The same pattern was observed regarding MPC. Conversely, Lin-/CD45-CD34+AC133+CXCR4+ cells appeared to circulate more in F-PMF, V617F-JAK2pos with a high IPSS (Table 1). No mutations were detected in the hot-spot of SRSF2 (codons 93-95) which have been associated to a fibrotic progression of PMF.
We also performed a bio-informatics analysis of microarray databases comparing the molecular libraries of the gene expression ommibus dataseries GSE3410 and GSE31869 (VSELs (Lin-CD45-CXCR4+CD34-) and CD34+ VSELs (Lin-CD45-CXCR4+CD34+) with the molecular profiling of circulating CD34+ cells from PMF patients and bone marrow CD34+ cells from healthy donors and we observed a tight clustering between PMF and VSELs cells samples (Figure 1).
Conclusions. Variations in the blood traffic of some progenitor cell subsets in patients with PMF were observed, including Lin-/CD45-CD34+AC133+CXCR4+ cells. The fact this cell subset was detected in higher number in F-PMF and in patients with a high IPSS, as well as a higher expression of OCT4 in F-PMF supports in part our hypothesis that PMF evolution can be associated to the recruitment and circulation of some primitive progenitors (dormant in the adult life) as it can be observed during the fetal period.

Figure 1 Figure 1.

Disclosures
No relevant conflicts of interest to declare.