Microarray Analysis of Gene Expression

Gene expression in the cells was determined by microarray analysis using the Affymetrix Human Gene 2.0 ST Array, according to the Affymetrix protocols (Santa Clara, CA, USA). Signal intensities were measured using a GeneChip Scanner3000 7G (Affymetrix) and converted to numerical data using the Affymetrix Expression Console software 1.3.1 (Affymetrix). To identify candidate genes of potential significance in U937, we applied a 1.2-fold change cutoff, since the combined responses of a group of genes acting in concert might affect the physiology of the cell, as previously described [19 (link)]. The digitized data were analyzed using GeneSpring GX 13.1.0 software (Agilent Technologies, Santa Clara, CA, USA). Genes whose expression changed significantly with treatment were subjected to functional analysis using Ingenuity Pathway Analysis software (IPA, Ingenuity Systems, QIAGEN, www.qiagen.com/ingenuity) [20 (link)]. For IPA analyses, we used two scores: an ‘enrichment’ score (Fisher’s exact test P-value) that measures the overlap of observed and predicted regulated gene sets, and a Z-score that assesses the match of observed and predicted upregulation/downregulation patterns [21 (link)]. The inclusion criteria for genes selected for the analysis was a fold change > ± .8 [22 (link)].