Differential expression and interaction of β-catenin/ICAT proteins in NSC67657 induced monocytic differentiation of HL60 cells / 中国药理学通报

ABSTRACT

Aim To analyze differential expression and interaction of β-catenin/ICAT proteins in HL60 cells when they were induced into monocytic differentiation, and to figure out the mechanism of NSC67657 in cellu-lar induction. Methods HL60 cells were treated by 10 μmol · L-1 NSC67657 , and cellular differentiation could be observed by cytochemical staining and flow cytometry. Then, RT-PCR and Western blot were em-ployed to determine the differential expression of β-catenin/ICAT genes and proteins. Co-immunoprecipi-tation assay was used to confirm the interaction of β-catenin/ICAT proteins, and laser co-focus light mi-croscopy technology was used to co-indentify proteins differential expression and intracellular location. Re-sults HL60 cells could be induced into monocytic dif-ferentiation after 5 days treatment using 10μM NSC67657 . The CD14 ( +)% cells could be up to o-ver 90%, and cytochemical staining reports were con-sistent with this result. The expressions of ICAT gene and protein were up-regulated significantly ( P <0. 01 ) , but the expressions ofβ-catenin gene and pro-tein, on the contrary, were down-regulated(P<0. 05) when HL60 cells were induced into monocytic differen-tiation. From co-immunoprecipitation assay findings, ICAT protein interacted with β-catenin protein, and the absorbance of protein electrophoresis bands in-creased in differentiated cells. From laser co-focus light microscopy assay findings, the fluorescence of ICAT and β-catenin protein could be both observed in cytoplasm and nucleus. In drug treated HL60 cells, the fluorescence of ICAT protein was enhanced both in cytoplasm and nucleus, however, the fluorescence ofβ-catenin protein, which looked like transferring into different organelles, decreased significantly in nucleus, but increased in cytoplasm. Conclusions HL60 cells could be induced into monocytic differentiation by NSC67657 and β-catenin/ICAT proteins differentially expressed during cellular differentiation. The enhanced interaction of β-catenin/ICAT proteins and β-catenin protein transferring from nucleus into cytoplasm indi-cates that NSC67657 probably induces HL60 cells into monocytic differentiation through down-regulating β-catenin protein and blockingβ-catenin protein from nu-cleus.