Effects Of Natural Plant Drug IHA-01on The Microtubule And The Mechanism Research

Objective:To explore the effect of the nature plant drug IHA-01on tubulin depolymerization activity in vitro and the effect on tubulin of cancer cell, and explore the possible anti-tumor mechanisms.Methods:3(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT) was used to investigated the proliferation of GLC cells treated with different concentrations of IHA-01. The cell cycle distribution of GLC was examined by flow cytometry after coculture with IHA-01. Cellular microtubules after IHA-01treatment were observed with fluorescence microscopy. Western blot was performed to detected the expression of tubulin protein after treatment IHA-01. Extracting tubulin from fresh pig brain tissue by regulating the temperature two circulations in vitro and ultracentrifuge. Establishing the polymerization-depolymerization balanced system of tubulin in vitro and measure the absorbance value of tubulin to test the role of drug to tubulin.Results:IHA-01inhibited the growth of GLC cells in a concentration-dependent manner. Flow cotometry analysis revealed a significant increased concentration dependent accumulation of GLC cells in the G0/G1phase(75.02±2.35)%and campared with normal control group(49.66±1.34)%the difference is statistically significant. Immunocytochemistry revealed changes in the cellular microtubule network and formation of polymerized microtubules in IHA-01(4μmol/L)-treatd cells and for the performance of the microtube to gather around nuclear and cytoplasmic microtubules were reduced.The western blot analysis showed that the expression of tubulin protein were depressed effectively after treatment IHA-014μmol/L、8μmol/L. The IHA-01can restrain the absorbance value of tubuin in polymerization process. Concentration of2μmol/L、4μmol/L、8μmol/L of IHA-01can inhibits the riseing of absorbance values in the process of warming and with the increase of drug concentration in the maximum absorbance by the normal control group fell to0.158、0.132、0.098.Conclusion:IHA-01can inhibit cell cycle of tumor cells and inhibiting tumor cell microtubule polymerization assembly, and at the same time affect the expression of microtubule protein. On the role of the microtubules polymerization inhibition could be one of IHA-01antitumor mechanism.