Antiproliferative effect of indomethacin on CML cells is related to the suppression of STATs/Bcl-X_L signal pathway

Indomethacin (IN) can inhibit cyclooxygenase activity and is considered to exert antitumor action in a variety of cancer cells. In the present study, we investigated the underlying mechanism of its antiproliferative effect on chronic myeloid leukemia (CML) cells. We studied the role of signal transducer and activator of transcription 1 or 5 (STAT₁ or STAT₅) and Bcl-X_L proteins in IN-induced proliferative inhibition on CML cells. Both K562 cells and fresh bone marrow mononuclear cells from five CML patients were exposed to IN. Cell proliferation was determined by MTT assay. The expression of JAK₂, STAT₁, STAT₅, and Bcl-X_L proteins was probed with Western blotting. The level of phosphorylated STAT₁ (p-STAT₁) or STAT₅ (p-STAT₅) proteins was determined by coimmunoprecipitation combined with Western blotting. Intracellular localizations of both STAT₁/STAT₅ and p-STAT₁/p-STAT₅ were observed by indirect immunofluorescence assay. Our results showed that IN could inhibit the proliferation of CML cells in a dose-dependent manner (36–288 μg/ml). The expression of STAT₁ and STAT₅ was suppressed by IN both in a concentration-dependent manner and a time-dependent (0–36 h) manner. The levels of p-STAT₁ and p-STAT₅ were down-regulated by IN. A similar result was obtained for Bcl-X_L protein expression. The intracellular fluorescence signals representing STAT₁/STAT₅ and p-STAT₁/p-STAT₅ were obviously weakened by IN. In contrast with IN, granulocyte-macrophage colony-stimulating factor could significantly promote the growth of CML cells and up-regulate the expression of both STAT₁/STAT₅ and p-STAT₁/p-STAT₅. This data indicated that IN is able to suppress the proliferation of CML cells, and the mechanism is associated with the inhibition of STATs/ Bcl-X_L signal transduction pathway.