A Sensitive Cell Screening Of Apoptosis Induced By Deoxynivalenol

Deoxynivalenol (DON) occours worldwidly in cereal grains and their feeds, which have been associated with human and animal toxicoses, serious questions remain regarding how it should be regulate. We adopted cell lines from different origin, and compared their sensitivity about DON from cytotoxicity, morphology and early apoptosis. The aim of this study was to establish fundament for the further mechanism investigation of apoptosis induced by DON.TEST 1:In the present experiment, cell lines from different origins (BHK-21,PIEC,HT-29,CHO-K1,BGC-823,BT-325 and HL-7702) were exposed to five different concentration of DON (125 ng/mL, 250 ng/mL, 500 ng/mL, 1000 ng/mL, 2000 ng/mL) for 24 h, 48 h and 72 h, respectively. The MTT bioassay was performed to determine the inhibition ratio on proliferation of the seven cell lines, and compared their sensitivity. The results showed that the different concentrations of DON could inhibit the proliferation of these cell lines. High inhibition depended on the increased concentration. The inhibition on proliferation of each concentation was the extremely significant compared to the controls (P<0.01). Significant time-dependent manner(P<0.05) was observerd in DON 500～2000 ng/mL groups of these cell lines from 24 h, 48 h and 72 h, except CHO-K1 in 48 h and 72 h groups. There was no significant change of inhibition in DON 1000 and 2000ng/mL groups in each time except BGC-823 and BT-325. The IC₅₀ value of BHK-21 was 333 ng/mL, after 24 h exposured to DON, and the others exceeded 2000ng/mL. The sensitivity of the cell lines were found in the decreasing order of BHK-21 > PIEC > HT-29 > CHO-K1> BGC-823 > BT-325 > HL-7702 after 48-h exposure, with IC₅₀ values of 320, 361, 598, 620, 1048, 1485, >2000ng/mL respectively. The sensitivities of the cell lines were found to be in the decreasing order of BHK-21 > PIEC > HT-29> CHO-K1> BT-325 > BGC-823> HL-7702 after 72-h exposure, with IC₅₀ values of 228, 238, 438, 678, 832, 908, >2000ng/mL, respectively. In conclusion, DON was toxical effect for these cell lines. The cytotoxicity was associated with does, duration time, cell origin and proliferation velocity. BHK-21 was the most sensitive about DON in these cells. However, HL-7702 was the less sensitive about DON than others.TEST 2:In the present experiment, cell lines from different origins (BHK-21,PIEC,HT-29,CHO-K1,BGC-823,BT-325 and HL-7702) were exposed to five different concentrations of DON (125 ng/mL, 250 ng/mL, 500 ng/mL, 1000 ng/mL, 2000ng/mL) for 24 h. The morphological changes of the cell lines were observed under microscope after staining. The apoptotic ratio was analyzed by flow cytometry to compare their sensitivity about DON. The DNA damage of sensitive cell was detected by comet assay. The result showed that Different concentration of DON induced apoptosis of all the cell lines The morphological changes and the apoptotic ratio of the cell lines was observed in a significant dose-dependent manner. A significant change of apoptotic ratio of these cell lines in defferent concentrations comparing with their control group(P<0.05), except BGC-823 in DON 125 ng/mL. The necrosis cell were less than 5% of the total cell numbers. The apoptotic ratio reached the highest level at 2000 ng/mL in these cell lines. In conclusion, DON can induced apoptosis in these cell lines. The morphological changes and the apoptotic ratio of the cell lines were associated with does, cell origin and cytotoxicity. BHK-21 was the most sensitive for DON and HL-7702 is the less sensitive about DON than others. The sensitivities of the cell lines were found to be in the decreasing order of HT-29>BHK-21>BT-325>PIEC > BGC-823, CHO-K1, HL-7702. The effect of DON on DNA damage of HT-29 is associated with apoptosis. HT-29 could adopt for the further mechanism investigation of apoptosis induced by DON.