Effect Of Arsenic Trioxide On VEGF/R And MMP-2,9 Expressed In K562 Cell

Objective:To investigate the effect of arsenic trioxide (ATO) on K562 cells, we detected the expression of VEGF, VEGFR, and the activity of MMP-2, 9 in K562 cells treated with ATO, further elucidating the possible mechanism of anti-leukemic angiogenesis by ATO and providing experimental data for clinic applying ATO to treat leukemia. Subject:Human leukemia cell line: K562 Methods:The ATO concentration was set as follows: 0, 0.05, 0.1, 0.2, 0.4, 0.8, 1.6, 3.2 and 6.4μM. K562 cells were seeded in 96-well plates (2×104 · well-1) and harvested after 72h incubation with indicated ATO. The cytotoxicity of ATO was detected by MTT assay (denoted with half-inhibition ratio IC50). According to the result of MTT, we selected 0, 0.05, 0.4 and 3.2μM ATO to do further experiment. After cells were seeded 24, 48 and 72h, the level of VEGF in supernatant was detected by enzyme-linked immunosorbent assay (ELISA), the expression ratio of VEGFR on cells were determined by FCM and the activity of MMP-2, 9 were assessed by gelatin zymography assay. Results:1. The IC50 of K562 was 2.12±0.11μM( x ±s) detected with MTT assay. Treated with 0.05μM ATO, proliferation of K562 cells were slightly up-regulated (P>0.05).Compared to control group, proliferation of the cells showed significant inhibition at the concentration of 0.4-6.4μM ATO (P<0.05); while mild inhibition was seen at 0.1-0.2μM ATO (but no significant decrease, P>0.05).2. The expression of VEGF was influenced by ATO. The expression of VEGF was prominently inhibited by 0.4μM and 3.2μM ATO (P<0.05) and slightly up-regulated by 0.05μM ATO (P>0.05). ATO had no influence on VEGFR.3. K562 cells expressed both MMP-2 and MMP-9. Among the three kinds of strips in the control group, the electrophoresis strip of MMP-2 (72KD) was the widest. The activity of MMP-2, 9 was partly inhibited by ATO. With the increase of ATO concentration and the incubation time, the inhibition was reinforced.Conclusions:1. 0.4-6.4μM ATO can inhibit the proliferation of K562 cells.2. 0.4 and 3.2μM ATO can inhibit the expression of VEGF and the activity of MMP-2, 9, but it doesn't influence the expression of VEGFR.3. The possible mechanism of anti-leukemic angiogenesis by ATO may be achieved by inhibiting the expression of VEGF and MMP-2, 9.