The G-CSF Produces Long-Term Changes in Gene and Mirnas Expression Profiles in CD34+ From Healthy Donors

The G-CSF (granulocyte colony-stimulating factor) is the cytokine most commonly used for the mobilization of hematopoietic progenitor cells (HPCs) from healthy donors used in allogenic transplantation (allo-HSCT). Although the administration of G-CSF is considered safe, the knowledge about its long-term effects, especially in HPCs, is limited. The aim of this study was to analyze whether G-CSF induces changes in gene and miRNAs expression profiles in HPCs from healthy donors, and determine whether or not these changes persist at the long term.

CD34 + cells were isolated by immunomagnetic separation and sorting from 5 healthy donors before mobilization with G-CSF and at afterwards at 5, 30 and 365 days after mobilization. A pool of samples from PB not mobilized was used as reference group. We analyzed the expression of 375 miRNAs using TaqMan MicroRNA Arrays Human v2.0 (Applied Biosystems), and the gene expression profile using Whole Human Genome Oligo microarray kit 4×44K (Agilent). The expression levels of genes and miRNAs were obtained by the 2^-ΔΔCTmethod. From expression data hierarchical clustering was performed using the Euclidean distance. To identify genes and miRNAs differentially expressed due to the effect of G-CSF at different time-points a cut-off value of level expression 2,5 above or below the control values was used and non-parametric Mann-Whitney test was applied. All analysis were performed using the Multi-experiment Viewer 4.7.1. The function of the miRNAs and genes of interest was determined from the various databases available online (TAM database, Gene Ontology, TargetScan Human).

Seven miRNAs were differentially expressed in HPCs at 5, 30 and 365 days after mobilization with G-CSF, as compared to HPCs obtained from not mobilized PB. In addition, 67 genes were also differentially expressed after administration of G-CSF, whose expression profiles remained abnormal 1 year after mobilization.

These genes are involved in biological processes such as hematopoietic cell proliferation, ribosomal protein synthesis, cell metabolism and transmembrane transportation. Interestingly, 8 of these genes are target of the miRNAs also identified in the current study, which suggests that their expression might be regulated by these miRNAs.

The G-CSF modifies gene expression profiles and miRNAs in HPCs from healthy donors. Remarkably, these changes were observed from early time-points and persisted at least 1 year after exposure to the drug. This effect of G-CSF on HPCs have not been previously reported and might be clinically relevant.

No relevant conflicts of interest to declare.