ICAT as a potential enhancer of monocytic differentiation: implications from the comparative proteome analysis of the HL60 cell line stimulated by all-trans retinoic acid and NSC67657.

A novel sterol mesylate compound (NSC67657) was recently identified and reported by National Cancer Institute that could efficiently induce the differentiation of HL60 cells into monocytes in vitro and in vivo. The expression of many proteins would have been changed during the differentiation process, and some proteins may have played key roles in the differentiation of HL60 cell line induced by this drug. Therefore, we treated HL60 cells with NSC67657 and all-trans retinoic acid (ATRA) to identify the differentially expressed proteins and determine their functions in cellular differentiation. Of the 45 differentially expressed protein spots investigated, 24 were either elevated or decreased in both the monocytic and granulocytic differentiating HL60 cells, 8 showed significant changes only when induced by NSC67657, and 13 showed significant changes only when induced by ATRA. After verification by RT-PCR, Western blotting, and immunocytochemistry, only the protein ICAT was found to be elevated by NSC67657 treatment alone. Although the over-expression of ICAT is not sufficient to induce the differentiation of HL60 cells into monocytes, it did increase the proportion of CD14+ cells in cells pretreated with NSC67657. Successful application of multiple techniques including two-dimensional gel electrophoresis, matrix-assisted laser desorption ionization time-of-flight mass spectrometry, Western blotting, and eukaryotic electroporation revealed that proteomic and molecular biological analyses provide valuable tools in drug development research.

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