Effect Of Evodiamine On The Microtubule In SPC-A1Cell

Evodia rutaecarpa is the dried and ripened fruits of Evodia rutaecarpa (Juss.)Benth,Evodia rutaecarpa (Juss.) Benth. var. officinalis (Dode) Huang or Evodiarutaecarpa (Juss.) Benth. var. bodinieri (Dode) Huang. Evodiamine had significantanti-tumor activity whatever it was in vivo or in vitro. Evodiamine can induceSPC-A1cells apoptosis through caspase-3,8-dependent pathway, which is related toG2/M arrest of the cell cycle.To explore the effect of evodiamine on the microtubule in SPC-A1cell. Theexpression and distribution of α-tubulin in SPC-A1cells were investigated byconfocal microscopy after staining with Anti-α-tubulin mouse monoclonalantibody.The inhibition of evodiamine in the presence or absence of MG132onSPC-A1cells was determined by MTT [3-(4,5-dimethylt hiazol-2-yl)-2,5-diphenylte-trazolium bromide] assay. The protein expression levels of α-，β-，γ-tubulin associatedwith microtubule of SPC-A1cells treated with evodiamine in the presence or absenceof MG132were measured by Western blots. The effects of evodiamine in the presenceor absence of MG132on the cell cycle distribution were determined using propidiumiodide staining and through flow cytometry. It showed an abnormal microtubulincytoskeleton morphology in SPC-A1treated with different concentration or time point（0,0.25,0.5,0.75,1μg/ml for4h or0.5μg/ml for8h）by confocal microscopy.Thefracture of microtubulin cytoskeleton morphology became more and more serious in adose-and time-dependent manner. Evodiamine had great inhibitory effects on theproliferation in SPC-A1cells, while MG132had no effects on the proliferation inSPC-A1cells. In the presence of MG132, the inhibition on the proliferation inSPC-A1cells caused by evodiamine were reduced. Evodiamine inhibited theexpression of α-tubulin, while had no effect on β-, γ-tubulin in SPC-A1cells.Thesuppressive effect of evodiamine on α-tubulin was abrogated by MG132. It showed asignificant increase in G2/M phase with the treatment of evodiamine in SPC-A1cells. The accumulation of cells at G2/M-phase was accompanied by a reduction ofcells at G0/G1phase. It suggested the cells were blocked at G2/M phase,the effectalso be abrogated by MG132. Evodiamine had significant inhibition in SPC-A1cells which is related to G2/Marrest of the cell cycle by the post-transcriptional control of α-tubulin.