Gene expression profiling during all-trans retinoic acid-induced cell differentiation of acute promyelocytic leukemia cells

Abstract

Using cDNA microarrays we determined the gene expression patterns in the human acute promyelocytic leukemia (APL) cell line NB4 during all-trans retinoic acid (ATRA)-induced differentiation. We analyzed the expression of 12,288 genes in the NB4 cells after 12 hours, 24 hours, 48 hours, 72 hours, and 96 hours of ATRA exposure. During this time course, we found 168 up-regulated and more than 179 down-regulated genes, most of which have not been reported before. Many of the altered genes encode products that participate in signaling pathways, cell differentiation, programmed cell death, transcription regulation, and production of cytokines and chemokines. Of interest, the CD52 and protein kinase A regulatory subunit alpha (PKA-Rlalpha) genes, whose products are being used as therapeutic targets for certain human neoplasias in currently ongoing clinical trials, were among the genes observed to be markedly up-regulated after ATRA treatment. The present study provides valuable data to further understand the mechanism of ATRA-induced APL cell differentiation and suggests potential therapeutic alternatives for this leukemia.

Figure 1.

Expression profiles of 12,288 genes in NB4 cells with or without ATRA (1 μmol/L) treatment. A: Hierarchical clustering of 12,288 genes based on their expression profiles in NB4 cells treated with ATRA at 12 hours, 24 hours, 48 hours, 72 hours, and 96 hours. Each row represents a single gene on the microarray, and each column a separated sample at a different time point. The expression profile of functionally related genes: mitogen activated protein kinase (B), protein kinase C and protein kinase A-related genes (C), IFN-induced genes (D), and neutrophil marker protein genes (E).

Figure 2.

RT-PCR analysis of genes in NB4 cells treated with ATRA. cDNA was synthesized from equal amounts of RNA isolated from NB4 cells treated with ATRA at 0, 12, 24, 48, 72, and 96 hours separately. The data are representative examples of three independent experiments that gave similar results. Actin serves as internal control.

Figure 3.

Differentiation of ATRA-induced NB4 cells. A: Morphological features of NB4 cells in response to ATRA treatment at different times. B: Nitroblue tetrazolium enzymatic reduction in ATRA-treated NB4 cells. C: CD11b intergrin expression following ATRA treatment of NB4 cells at each time point as assessed by flow cytometric analysis.