Figures & data
Figure 1 Methyl jasmonate (MeJ) suppressed the proliferation of KY170R, sh-KY170R, and scr-KY170R cells independent of the expression of AKR1C3.
Notes: (A) The construction of stable low expression of AKR1C3 in KY170R cells (sh-KY170R). Western blot (left) and real-time RT-PCR (right) showed AKR1C3 expression was lower in sh-KY170R cells. (B) The proliferation of KY170R, sh-KY170R, and scr-KY170R cells was analyzed by Alamar Blue assay after incubation with different concentrations of MeJ for 24, 48, or 72 h. There were no significant differences between the three cells (P>0.05). (C) Incubation of KY170R, sh-KY170R, and scr-KY170R cells with indicated concentrations of MeJ for 24 h; the inhibition rates of the three cell lines showed no differences (P>0.05). (D) The colony formation of KY170R, sh-KY170R, and scr-KY170R cells were counted after cells were treated with the indicated concentrations of MeJ for 7 days. A concentration of 200 μmol/L of MeJ had little effect on the growths of the three cells.
Abbreviations: AKR1C3, aldo-keto reductase family 1 member 3; MeJ, methyl jasmonate; RT-PCR, reverse-transcription PCR.
Figure 2 Radiosensitivity effect of MeJ depended on the expression of AKR1C3.
Notes: (A) KY170R cells (highly expressing AKR1C3) incubated with 200 μmol/L MeJ for 24 h before radiation were more sensitive to radiation than their untreated counterparts. MeJ could enhance the radiation sensitivity of KY170R cells that highly expressed AKR1C3. (B) sh-KY170R cells (not or lowly expressing AKR1C3) incubated with 200 μmol/L MeJ for 24 h were equally resistant to radiation as their untreated counterparts. (C) The survival curves of sh-KY170R and KY170R cells incubated with or without MeJ.
Abbreviations: AKR1C3, aldo-keto reductase family 1 member 3; IR, irradiation; MeJ, methyl jasmonate.
Figure 3 Methyl jasmonate (MeJ) could inhibit AKR1C3 enzyme expression and the 11-ketoprostaglandin reductase activity of AKR1C3.
Notes: (A) Western blots showed MeJ could reduce AKR1C3 protein expression in KY170R cells in a dose-dependent manner (indocin was a positive drug). (B) ELISA results showed that MeJ could increase the levels of PGD2, while decreasing the levels of PGF2. MeJ could inhibit the 11-ketoprostaglandin reductase activity of AKR1C3 in a dose-dependent manner; *P<0.05.
Abbreviations: AKR1C3, aldo-keto reductase family 1 member 3; PG, prostaglandin; ELISA, enzyme-linked immunosorbent assay; MeJ, methyl jasmonate.
Figure 4 Radiation sensitivity effects of MeJ depended on activation of the PPARγ pathway.
Notes: (A) KY170R cells incubated with different densities (0,100, and 200 μmol/L) of MeJ for 24 h. Expression levels of PPARγ in KY170R cells were detected by Western blot analysis. MeJ could activate the PPARγ pathyway. (B) Clone formation assays showed GW9662 can weaken the radiation sensitivity effects of MeJ on KY170R cells. The cloning numbers were significantly higher after cells were incubated with both MeJ and GW9662. (C) Surviving fraction results showed that cells incubated with GW9662 were no longer sensitive to the effects of MeJ (P>0.05); **P<0.01.
Abbreviations: PPARγ, peroxisome proliferator-activated receptor gamma; IR, irradiation; MeJ, methyl jasmonate.
Figure 5 The distribution of apoptotic KY170R cells before and after radiation.
Notes: (A) The cell apoptosis analysis of KY170R cells treated before radiation (n=3). Apoptotic ratio of cells increased when MeJ density increased (from 0 to 5000 μmol/L). Apoptotic rates between 200 µmol/L and control group were not significantly different (P>0.05). (B) The cell apoptosis analysis of KY170R cells treated 48 h after radiation (n=3). Apoptotic rates between drug group and control group were not significantly different (P>0.05). (C) Radiation of 4 Gy could increase the apoptotic rate of cells, while cells pretreated with 200 μmol/L MeJ did not exhibit an increase in the apoptotic rate (P>0.05); *P<0.05.
Abbreviations: MeJ, methyl jasmonate; IR, irradiation.
Figure 6 Radiation sensitivity effects of methyl jasmonate depended on the generation of ROS in KY170R cells.
Notes: (A) MeJ could increase the ROS levels in KY170R cells both before and after radiation (P>0.05). (B, C) Clone formation assays of KY170R cells. The ROS scavenger NAC could reverse the radiosensitivity effects of MeJ (P>0.05); **P<0.001.
Abbreviations: ROS, reactive oxygen species; NAC, N-acetyl cysteine; IR; MeJ, methyl jasmonate.
