| objective:To investigate the effect on the proliferation, apoptosis and cell cycle of Chinese medicine Art on human oral squamous carcinoma Tca8113 cell line, and test regulating function of tumor suppressor gene RECK protein expression. This study is provided impossible action mechanism of Art and the foundation theory for ART using in oral squamous carcinoma clinical treatment.Methods:Human oral squamous carcinoma Tca8113 cells are randomized into control,12.5ug/ml,25ug/ml,50ug/ml, 100ug/ml,200ug/ml groups, respectively cultivated for 24h,48h,72h:(1) Inverted microscope observes morphological changes of various concentration Art on Tca8113 cells; (2) MTT method observes the growth inhibition of various concentration Art on Tca8113 cells; (3) Flow cytometry analyzes the influence of apoptosis and cell cycle distribution of various concentration Art on Tca8113 cells; (4) Imm--unocyto chemistry method detect the influence of RECK protein expression of various concentration Art on Tca8113 cells. Results:(1) The Tca8113 cells grow border and clear on each time control group, show epithelioid macro--phages form, the nucleus is located in cell central; after 24h,12.5ug/ml,25ug/ml,50ug/ml concentration group cells form do not change; in 100ug/ml,200ug/ml concentration group, some cells go round, shrink, refractive sex is strong; After 48h and 72h, the cells present irregular form, and have large vacuoles appeared in shape, the burr on the edge, especially in refractive sex, 100ug/ml,200ug/ml concentration group is most obvious. (2) ART can inhibit the proliferation of human oral squamous carcinoma Tca8113 cell, and with the extension of time, the increase of the concentration, the inhibition is more obvious. At the same period, cell inhibition rate in various concentration groups compared with control groups has significantly increased, the difference is statistically significant (P<0.05); cell inhibition rate in the same concentration at 24h,48h,72h compared, the difference is statistically significant (P<0.05), and presents time and dose dependent, especially in the 100ug/ml,200ug/ml group, at 48h,72h, most obvious; cell inhibition rate is 25.74%,29.34%,33.25%,45.47% respectively. (3) ART can induce Tca8113 induced apoptosis of oral squamous cell carcinoma, and with the extension of role time, the increase of the concentration, the apoptotic cells is more increasing. At the same period, cell apoptosis rate in various concentration groups compared with control groups has significantly increased, the difference is statistically significant (P<0.05); cell apoptosis rate in the same concentration at 24h,48h,72h compared, the difference is statistically significant (P<0.05), and presents time and dose dependent, especially in the 100ug/ml,200ug/ml group, at 48h,72h, most obvious; cell apoptosis rate is (30.88±0.53)%, (37.86±0.49)%, (63.20±0.32)%, (83.43±0.44)% respectively. (4) After 24h,48h,72h, cell cycle distribution in various concentration groups changes, main show is that G0/G1 phase cell ratio increase, and G2/M phase, S phase cell ratio decrease. At the same period, G0/G1 phase cell ratio in various concentration groups compared with control groups has significantly increased, the difference is statistically significant (P<0.05); especially in the 100ug/ml,200ug/ml group, G0/G1 phase cell ratio is (66.92±0.53)%, (68.87±0.49)%, (67.98±0.57)%, (70.92±0.44)%, (72.95±0.68) %, (78.05±0.42)%respectively. Cell cycle distributionin in the same concentration at 24h,48h,72h compared, the difference is not statistically significant (P>0.05). (5) The result of immunocyto chemistry shows, After 24h, each average light density value of the RECK expression cell in various concentration groups compared with control groups has increased, but the difference is not statistically significant (P>0.05); After 48h,72h, each average light density value of the RECK expression cell in various concentration groups compared with control groups has significantly increased, the difference is statistically significant (P<0.05). In different period, average light density value in 12.5 ug/ml,25ug/ml group at 24h,48h,72h compared, the difference is not statistically significant (P>0.05); average light density value in 50ug/ml concentration group at 24h,48h,72h compared, the difference is statistically significant (P<0.05), and presents time, dose dependent in high concentration groups. Conclusion:1.Art can effectively inhibit the proliferation of Tca8113 cells in vitro in dose and time dependent manners.2. Art can induce the apoptosia of Tca8113 cells in dose and time dependent manners.3. Art can make the Tca8113 cell stuck in G0/G1 phase in dose-dependent manners, but in no time-dependent. 4. Art can obviously increase the expression of RECK protein and present time and dose dependent in high concentration groups.5. Chinese medicine Art has proliferation inhibition, retardents the cell cycle, induces the apoptosis, increase the expression of RECK protein, exerts its inhibiting tumor invasion and metastasis, which is prompted that Art in resistance to oral squamous cell carcinoma has potential value of basic and clinical research, and Art is expected to become the risk-benefit ratio anti-tumor and adjuvant chemotherapy new drug, further explore the mechanism of oral squamous cell carcinoma, improve the curative effect. |
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