Journal: Scientific Reports

Article Title: Indibulin dampens microtubule dynamics and produces synergistic antiproliferative effect with vinblastine in MCF-7 cells: Implications in cancer chemotherapy

doi: 10.1038/s41598-018-30376-y

Figure Lengend Snippet: Indibulin inhibited the proliferation of MCF-7 cells and blocked the cell cycle progression at mitosis: ( A ) Structure of indibulin. ( B ) MCF-7 cells were treated with a vehicle or increasing concentrations of indibulin for 48 h. The inhibition of cell proliferation was determined by the SRB assay. Data are average of three independent experiments. ( C ) Cells were treated with vehicle or different concentrations (300 and 600 nM) of indibulin for 48 h and stained with propidium iodide. The DNA content of the cells was quantified by a flow cytometer and the data were analyzed using the Modfit LT program (Verity Software, ME, USA). The dark black lines show the fitting of the data by Modfit LT program and in each panel, the peaks correspond to G1 phase (left-side red peak), S phase (middle hashed line peak) and G2/M phase (right-side red peak). ( D ) Cells were treated with a vehicle or different concentrations of indibulin for 48 h and then stained with an antibody against phosphohistone H3 (S10) (green), a mitotic marker. DNA stained with Hoechst is shown in blue. Scale bar = 10 µm. ( E ) The histogram shows the percentage of phosphohistone H3 (S10) positive cells in the presence of a vehicle or different concentrations of indibulin. Three hundred cells were counted using Hoechst staining. Data are average of three independent experiments and error bar represents S.D.

Article Snippet: Indibulin was purchased from Tocris Biosciences.

Techniques: Inhibition, Sulforhodamine B Assay, Staining, Flow Cytometry, Software, Marker

Journal: Scientific Reports

Article Title: Indibulin dampens microtubule dynamics and produces synergistic antiproliferative effect with vinblastine in MCF-7 cells: Implications in cancer chemotherapy

doi: 10.1038/s41598-018-30376-y

Figure Lengend Snippet: Effects of indibulin on interphase microtubules in MCF-7 cells: ( A ) Cells were incubated with vehicle or different concentrations of indibulin for 24 h and were then stained with an antibody against α-tubulin (red). DNA stained with Hoechst is shown in blue. Scale bar = 10 µm. ( B ) MCF-7 cells were treated with vehicle or 150, 450 and 900 nM indibulin (lanes 1–4, respectively) for 48 h. 25 nM vinblastine (lane 5) was used under similar experimental conditions as a control. Polymeric and soluble tubulin fractions were isolated, loaded separately on two different SDS-PAGEs and immunoblotted with the α-tubulin antibody.

Article Snippet: Indibulin was purchased from Tocris Biosciences.

Techniques: Incubation, Staining, Control, Isolation

Journal: Scientific Reports

Article Title: Indibulin dampens microtubule dynamics and produces synergistic antiproliferative effect with vinblastine in MCF-7 cells: Implications in cancer chemotherapy

doi: 10.1038/s41598-018-30376-y

Figure Lengend Snippet: Effects of indibulin on mitotic MCF-7 cells: Cells were treated without and with different concentrations of indibulin and were stained with antibodies against α-tubulin (red) and γ-tubulin (green). Scale bar = 10 µm.

Article Snippet: Indibulin was purchased from Tocris Biosciences.

Techniques: Staining

Journal: Scientific Reports

Article Title: Indibulin dampens microtubule dynamics and produces synergistic antiproliferative effect with vinblastine in MCF-7 cells: Implications in cancer chemotherapy

doi: 10.1038/s41598-018-30376-y

Figure Lengend Snippet: Indibulin suppressed microtubule dynamics in live MCF-7 cells: ( A ) Life history traces of microtubules of cells treated with (i) vehicle, (ii) 75 and (iii) 150 nM indibulin for 3 h. ( B ) Cells were treated with a vehicle or different concentrations (150 and 300 nM) of indibulin for 24 h and were processed for immunostaining with antibodies against EB1 (red) and α-tubulin (green). Boxed regions of merged panels are shown at higher magnification. Scale bar = 10 µm.

Article Snippet: Indibulin was purchased from Tocris Biosciences.

Techniques: Immunostaining

Journal: Scientific Reports

Article Title: Indibulin dampens microtubule dynamics and produces synergistic antiproliferative effect with vinblastine in MCF-7 cells: Implications in cancer chemotherapy

doi: 10.1038/s41598-018-30376-y

Figure Lengend Snippet: Indibulin affected the parameters of dynamic instability of interphase microtubules in MCF-7 cells.

Article Snippet: Indibulin was purchased from Tocris Biosciences.

Techniques: Control

Journal: Scientific Reports

Article Title: Indibulin dampens microtubule dynamics and produces synergistic antiproliferative effect with vinblastine in MCF-7 cells: Implications in cancer chemotherapy

doi: 10.1038/s41598-018-30376-y

Figure Lengend Snippet: Indibulin activated the mitotic checkpoints in MCF-7 cells: ( A , B ) MCF-7 cells were incubated with a vehicle or different concentrations of indibulin for 24 h and processed for immunostaining with antibodies against Hec1 (red) and Mad2 (green) ( A ) or BubR1 (red) ( B ). Scale bar = 10 µm.

Article Snippet: Indibulin was purchased from Tocris Biosciences.

Techniques: Incubation, Immunostaining

Journal: Scientific Reports

Article Title: Indibulin dampens microtubule dynamics and produces synergistic antiproliferative effect with vinblastine in MCF-7 cells: Implications in cancer chemotherapy

doi: 10.1038/s41598-018-30376-y

Figure Lengend Snippet: Indibulin increased the level of Mad2 and BubR1 and activated apoptosis in MCF-7 cells: ( A ) MCF-7 cells were incubated without and with different concentrations of indibulin and the cell extract was prepared. Vinblastine (25 nM) was used as a positive control. Western blot was performed with the extracts and immunoblotting was done with anti-Mad2 IgG, anti-BubR1 IgG, and anti-β actin IgG. The appropriate molecular weight bands of Mad2 (24 kDa), BubR1 (120 kDa) and β-actin (42 kDa) were cut from the same gel and immunoblotted with respective antibodies. The experiment was performed three times, shown is one of the representative blots. ( B ) MCF-7 cells were incubated with vehicle or different concentrations of indibulin for 48 h and then stained with annexin V (green) and propidium iodide (red) for detecting apoptosis. Scale bar = 10 µm. ( C ) Western blot of MCF-7 whole cell extracts prepared after incubating cells without and with different concentrations of indibulin and vinblastine (25 nM) for 48 h. The cell extract was separated on SDS-PAGE and appropriate molecular bands of PARP (116 kDa) and β-actin (42 kDa) were cut from the same gel. Immunoblotting was done with anti-PARP IgG and anti-β actin IgG. Shown is one of the representative blots from three experiments.

Article Snippet: Indibulin was purchased from Tocris Biosciences.

Techniques: Incubation, Positive Control, Western Blot, Molecular Weight, Staining, SDS Page

Journal: Scientific Reports

Article Title: Indibulin dampens microtubule dynamics and produces synergistic antiproliferative effect with vinblastine in MCF-7 cells: Implications in cancer chemotherapy

doi: 10.1038/s41598-018-30376-y

Figure Lengend Snippet: Indibulin treatment induced apoptosis in MCF-7 cells.

Article Snippet: Indibulin was purchased from Tocris Biosciences.

Techniques: Control

Journal: Scientific Reports

Article Title: Indibulin dampens microtubule dynamics and produces synergistic antiproliferative effect with vinblastine in MCF-7 cells: Implications in cancer chemotherapy

doi: 10.1038/s41598-018-30376-y

Figure Lengend Snippet: Indibulin and vinblastine exert synergistic anti-proliferative effects on MCF-7 cells. The median dose plot for the inhibition of cell proliferation in the presence of indibulin (i) and vinblastine (ii) is shown. Histogram (iii) shows the combination indices for the combination of indibulin (IN) with vinblastine (VB). Data are average of three independent experiments and represent mean ± SD.

Article Snippet: Indibulin was purchased from Tocris Biosciences.

Techniques: Inhibition

Journal: Scientific Reports

Article Title: Indibulin dampens microtubule dynamics and produces synergistic antiproliferative effect with vinblastine in MCF-7 cells: Implications in cancer chemotherapy

doi: 10.1038/s41598-018-30376-y

Figure Lengend Snippet: Effects of indibulin on undifferentiated and differentiated SH-SY5Y cells. ( A ) Undifferentiated SH-SY5Y cells were incubated in the absence and presence of indibulin (25 nM), vinblastine (10 nM) and colchicine (10 nM) for 24 h. Cells were fixed and processed for immunostaining with antibodies against α-tubulin (green) and acetylated tubulin (red). Scale bar = 10 μm. ( B ) Differentiated SH-SY5Y cells were treated and processed as in ( A ) and observed at two magnifications under a microscope. Scale bar = 100 and 10 μm in low (Left panel) and high (Right panel) magnification, respectively.

Article Snippet: Indibulin was purchased from Tocris Biosciences.

Techniques: Incubation, Immunostaining, Microscopy