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To investigate the potential anti-proliferative and pro-apoptotic effects of 3,3-diindolylmethane(DIM), important derivatives of Indole-3-carbinol(I3C) on Siha cells.
Cell proliferation was detected by cell counting it-8 and cell apoptosis was analyzed by flow cytometry. Morphological changes accompanying cell apoptosis were observed with fluorescence micro-scope after Hoechst 33258 staining. In addition, changes of proteins expression involved in MPAK and PI3K pathway were determined by western blotting.
DIM treatment inhibited the proliferation and induced the apoptosis of SiHa cells significantly in a time-and dose-dependent manner. After DIM treatment for 48h, the calculated IC50 of DIM for Siha cells was 44.44umol/L. The apoptosis ratio in SiHa cells were(4.56±0.52)%,(10.09±1.32)%,(21.11±3.36)% and (55.56±6.33)% at 0,25,50 and 100umol/L DIM. Evident morphological changes including memebrane shrinking, round shaping and budding to form apoptotic bodies were observed in SiHa cells after treatment for 48h at different concentration. In addition, expressions of ERK, p-ERK, p38 and p-p38, which were involved in MAPK pathway, were down-regulated when the dose of DIM increased. Another proteins involved in MAPK pathway, such as JNK and p-JNK were up-regulated. Furthermore, DIM treatment significantly suppressed the expressions of Akt, p-Akt, PI3K, p110α, PI3K p110β,PI3K class III, GSK3-β,p-PDKI and p-c-Raf which were involved in PI3K Pathway.
These results demonstrated that DIM exerted anti-tumor effects on SiHa cells through its anti-proliferative and pro-apoptotic roles. The molecular mechanism for these effects may be related to its regulatory effects on MAPK and PI3K pathway and apoptosis proteins. DIM might be a preventive and therapeutic agent against cervical cancer.
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