Differential expression of multiple unexpected genes during U937 cell and macrophage differentiation detected by suppressive subtractive hybridization.

OBJECTIVE: The objective of this study was to identify new markers of myelomonocytic differentiation using a sensitive technique that permits detection of rare differential gene expression.MATERIALS AND METHODS: [corrected] Suppressive subtractive hybridization (SSH) was performed between the human myelomonocytic U937 cell line and 1 alpha, 25-dihydroxyvitamin D3 and transforming growth factor beta 1 differentiated U937 cells. cDNA clones with significant increased expression in differentiated U937 cells over nondifferentiated U937 cells were characterized by sequencing. [corrected] The pattern of differential gene expression obtained by SSH was confirmed by cDNA Southern and Northern blots on the undifferentiated vs. differentiated U937 cells, and by reverse transcriptase polymerase chain reaction on undifferentiated human CD34(+) stem cells isolated from bone marrow vs. peripheral blood CD14(+) mature monocytes.RESULTS: Seven cDNAs never associated with in vitro U937 cell myelomonocytic differentiation (prolactin, 11-beta hydroxysteroid dehydrogenase [11 beta-HSD)] haptoglobin alpha (2FS)-beta precursor, GLIPR, RTVP, the RNA helicase P68, and spermidine-spermine N1-acetyltransferase) were identified. The first five of these genes previously were associated with immune function and the last two are important for intermediary metabolism. Differential expression was confirmed in CD34(+)/CD14(+) monocyte differentiation for all genes but 11 beta-HSD.CONCLUSIONS: We identified six new markers of U937 cell differentiation, which also are differentially expressed during normal human myelomonocytic differentiation.