Characterization of gene expression of CD34+ cells from normal and myelodysplastic bone marrow

Abstract

Gene patterns of expression in purified CD34 ؉ bone marrow cells from 7 patients with low-risk myelodysplastic syndrome (MDS) and 4 patients with high-risk MDS were compared with expression data from CD34 ؉ bone marrow cells from 4 healthy control subjects. CD34 ؉ cells were isolated by magnetic cell separation, and high-density oligonucleotide microarray analysis was performed. For confirmation, the expression of selected genes was analyzed by real-time polymerase chain reaction. Class membership prediction analysis selected 11 genes. Using the expression profile of these genes, we were able to discriminate patients with low-risk from patients with high-risk MDS and both patient groups from the control group by hierarchical clustering (Spearman confidence). The power of these 11 genes was verified by applying the algorithm to an unknown test set containing expression data from 8 additional patients with MDS (3 at low risk, 5 at high risk). Patients at low risk could be distinguished from those at high risk by clustering analysis. In low-risk MDS, we found that the retinoic-acid-induced gene (RAI3), the radiation-inducible, immediateearly response gene (IEX1), and the stress-induced phosphoprotein 1 (STIP1) were down-regulated. These data suggest that CD34 ؉ cells from patients with low-risk MDS lack defensive proteins, resulting in their susceptibility to cell damage.

dlk1/FA1 Regulates the Function of Human Bone Marrow Mesenchymal Stem Cells by Modulating Gene Expression of Pro-inflammatory Cytokines and Immune Response-related Factors

“…Also, the analysis of hematopoiesis in Dlk1 knock-out mice suggested the contribution of dlk1 in lineage-specific development of megakaryocytes and B cells (8). In addition, Dlk1 was elevated by CD34ϩ cells in patients with low risk myelodysplastic syndrome and acute myeloid leukemia as compared with normal individuals (8,77). One of the main functions of MSC is to provide a hematopoiesis-supporting microenvironment via the expression of many cytokines and regulatory factors (78).…”

The NLRP3 inflammasome functions as a driver of the myelodysplastic syndrome phenotype

“…[10][11][12][13] Furthermore, MDS hematopoietic stem and progenitor cells (HSPCs) overexpress TLRs, which is accompanied by activation of respective signaling intermediates and has been implicated in the aberrant proliferation of HSPCs and the pathogenesis of peripheral blood cytopenias. [14][15][16]NF-kB-induced transcriptional priming of inflammasome proteins, followed by cation channel activation, cell volume expansion, and inflammasome component assembly. 17,18,20,21 NLRP3 is activated by diverse DAMP signals, including S100A9 homodimers and S100A8/9 heterodimers that function as alarmins that converge upon NADPH oxidase to generate reactive oxygen species (ROS).…”

Prediction of qualitative outcome of oligonucleotide microarray hybridization by measurement of RNA integrity using the 2100 Bioanalyzer™ capillary electrophoresis system

“…Due to the limited RNA content in certain sample preparations (e.g., CD34-selected cells), a double in vitro transcription technique (nanogram-scale assay) was used in more than half of the experiments (n=233, see Table 1). To assay 50-ng total RNA, the standard Affymetrix target amplification protocol was modified by using the firstround complementary RNA (cRNA) product to generate double-stranded complementary DNA that was then used for a second round of in vitro transcription for synthesis of biotinylated cRNA [9].…”