Abstract
BackgroundPrevious studies demonstrated that selenite induced cancer-cell apoptosis through multiple mechanisms; however, effects of selenite on microtubules in leukemic cells have not been demonstrated.MethodsThe toxic effect of selenite on leukemic HL60 cells was performed with cell counting kit 8. Selenite effects on cell cycle distribution and apoptosis induction were determined by flow cytometry. The contents of cyclin B1, Mcl-1, AIF, cytochrome C, insoluble and soluble tubulins were detected with western blotting. Microtubules were visualized with indirect immunofluorescence microscopy. The interaction between CDK1 and Mcl-1 was assessed with immunoprecipitation. Decreasing Mcl-1 and cyclin B1 expression were carried out through siRNA interference. The alterations of Mcl-1 and cyclin B1 in animal model were detected with either immunohistochemical staining or western blotting. In situ detection of apoptotic ratio was performed with TUNEL assay.ResultsOur current results showed that selenite inhibited the growth of HL60 cells and induced mitochondrial-related apoptosis. Furthermore, we found that microtubule assembly in HL60 cells was altered, those cells were arrested at G2/M phase, and Cyclin B1 was up-regulated and interacted with CDK1, which led to down-regulation of the anti-apoptotic protein Mcl-1. Finally, in vivo experiments confirmed the in vitro microtubule disruption effect and alterations in Cyclin B1 and Mcl-1 levels by selenite.ConclusionsTaken together, the results from our study indicate that microtubules are novel targets of selenite in leukemic HL60 cells.
Highlights
Microtubules have important roles in many cell behaviors such as cell division, organelle positioning, vesicular transport and cell-shape determination [1,2,3]
Taken together, the results from our study indicate that microtubules are novel targets of selenite in leukemic HL60 cells
To identify which pathway participated in selenite-induced apoptosis, we extracted the mitochondrial fraction of HL60 cells after they were exposed to selenite and discovered that cytochrome C and AIF were released (Figure 1C), which suggested that a mitochondrial-related apoptotic pathway was utilized
Summary
Microtubules have important roles in many cell behaviors such as cell division, organelle positioning, vesicular transport and cell-shape determination [1,2,3]. Previous studies have showed that microtubule dynamics are necessary for these functions in vivo [2,4,5,6]. Chemicals affecting microtubule dynamics often impact these functions in vivo. Many anti-tumor agents have been developed for their effects on microtubule dynamics and cell-cycle distribution [7,8,9,10,11,12]. Previous studies demonstrated that selenite induced cancer-cell apoptosis through multiple mechanisms; effects of selenite on microtubules in leukemic cells have not been demonstrated
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