Gene expression analysis of cord blood stem cells

Cord blood (CB) is an attractive source for hematopoietic cellreplacement and CD34+ and CD133+ cells are the most clinically relevantcell subsets used for transplantation purposes. In view of characterizing thegene profile of these two populations of hematopoietic stem cells (HSCs), agenome-wide gene expression analysis is required. A powerful tool tobroaden the knowledge of molecular mechanisms of HSCs is geneexpression analysis using microarray technology. In this study wemeasured gene expression profiles of CD34+ and CD133+ CB cells usingAffymetrix HG-U133A arrays containing 22.000 probes. We processed 10CB units: CD34+ (n=5) and CD133+ (n=5) cells were selected using theMiniMacs immunomagnetic separation system (Miltenyi Biotec). Weevaluated RNA quantity and quality using the RNA 6000 Nano LabChipkit (Agilent Technologies). Given the low amount of RNA due to thelimiting numbers of HSCs obtained, we used a double-cycle amplificationprotocol to obtain biotin-labeled target for hybridization starting from aslow as 100 ng of total RNA. Gene expression profiling data were analyzedby means of two different standard software packages (MAS 5.0 anddChip). A supervised analysis allowed us to identify genes distinguishingthe two different groups of samples, one containing the CB CD34+ and theother containing the CB CD133+ samples. The comparison showed theover-expression of genes involved in cell homing (IL-8, CXCL4, CXCL7,fibronectin 1, collagen type IV) and myeloid differentiation (GMCSFR,HDC, MNDA, MRP) in the CD34+ subset. Moreover, no gene was foundto be up-regulated in CD133+ subset. These preliminary data on the geneexpression profiles of these two populations support the concept thatCD34+ cells represent a more committed, hematopoietic cell subsetcompared to CD133+ cells. In conclusion, our study may serve as a basisfor further researches in the field of functional genomic of HSCs.