Mechanism of the inhibition of leukemia cell growth and induction of apoptosis through the activation of ATR and PTEN by the topoisomerase inhibitor 3EZ, 20Ac-ingenol.

The PI3K/Akt signaling pathway is constitutively activated in various leukemias. In the present study, the topoisomerase inhibitor, 3EZ, 20Ac-ingenol, was more effective in inhibiting the growth of BALL-1 cells than that of normal lymphocyte cells. ATM/ATR protein levels were increased, PTEN protein was upregulated, and p-Akt protein was downregulated at early time points after treatment with 3EZ, 20Ac-ingenol. In further experiments, p53 protein expression was increased, and H2AX phosphorylation and p21 protein expression were induced after treatment with 3EZ, 20Ac-ingenol. Moreover, the activation of caspase 3 followed decrease in the Bcl-2/Bax ratio after treatment with 3EZ, 20Ac-ingenol, and accumulation of sub-G1 phase cells was observed in flow cytometry analyses. These data suggest that 3EZ, 20Ac-ingenol-induced DNA damage downregulates p-Akt and upregulates ATR leading to cell cycle arrest and increased apoptosis in BALL-1 cells.

3EZ,20Ac-ingenol, a catalytic inhibitor of topoisomerases, downregulates p-Akt and induces DSBs and apoptosis of DT40 cells.

We have previously reported that many ingenol compounds derived from Euphorbia kansui exhibit topoisomerase (topo) II inhibitory activity. Of these compounds, 3EZ,20Ac-ingenol inhibited topo I activity. Camptothecin, which inhibits the religation activity of topo I without interfering with the binding of topo I to DNA and induces topo I-mediated DNA cleavage, was used as a positive control. In this study, we found that 3EZ,20Ac-ingenol did not hamper the binding of topo I to DNA in the same manner as camptothecin but affected the inhibition of cleavage of one DNA strand. 3EZ,20Ac-ingenol inhibited cell proliferation by blocking cell cycle progression in the G2/M phase. To define the mechanism of inhibition of DT40 cell proliferation, the change in Akt activity was observed because Akt activity is regulated in response to DNA damage. Western blot analysis revealed that 3EZ,20Ac-ingenol downregulated the expression of p-Akt, and apoptosis was detected by the presence of DNA double-strand breaks and caspase 3 activation.

Analysis of How a Biofilm Forms on the Surface of the Aquatic Macrophyte Phragmites australis.

The process by which a biofilm forms on the surface of the aquatic macrophyte Phragmites australis was investigated over a period of about two months (from mid-May to late-July, 2008) in Lake Biwa. The biofilm formed relatively quickly, its wet weight per unit area after seven day being that of a mature biofilm. This speed can be attributed to the many active bacteria in the early stage of its formation and the extracellular polymeric substances (EPS) they produce. The EPS carried electric charges that attracted nutrient ions from surrounding lake water, which, by electrostatic interaction, reached a high concentration as early as day 7 of the formation process. This significantly affected the biofilm community, which differed greatly from that of the lake water even at the beginning of biofilm formation. Brown amorphous compounds (a complex of organic and inorganic substances), covered the biofilm in the second half of its formation process producing a different community structure from that initially. This study revealed a fast and dynamic process of biofilm formation on the reed surface of reed.