| Objective: To investigate the effects of 5-allyl-7-gen-difluoromethyllenechrysin (ADFMChR), a novel chrysin derivative on the geowth inhibition and induction of apoptosis of human ovarian cancer(CoC1) cell line in vitro , and its mechanism whether involve activation of PPARγ.Methods: Human ovarian cancer (CoC1) cells were cultured in vitro. MTT assay was used to determine the effect of ADFMChR on the viability of CoC1 cells. The trypan blue exclusion method was used to count the number of viablity cells, to draw cell growth curve and to investigate the growth inhibitory effect of ADFMChR on CoC1 cells. Colony formation in soft agar was used to test inhibition of colony formation in CoC1 cells treated with ADFMChR. DNA agarose gel electrophoresis was used to test apotosis induced by ADFMChR in CoC1 cell line. Flow cytometry (FCM) using PI staining was used to analyse the apoptosis rate of CoC1 cells treated with ADFMChR. Western blot was used to analyze expression of PPARγ,NF-κB,Bcl-2,Bax protein in CoC1 cells .Results: The MTT assay showed that cell viability inhibitory rate was 26.4%,40.6%,51.3%,68.4%,75.8%和57.5%, respectively, after CoC1 cell treatment with ADFMChR at various concentration of 1.0,3.0,10.0,30.0,100.0μmol/L and ChR at 100.0μmol/L for 48h. its IC50 was 7.26μmol/L. The cell viability inhibitory rate of ADFMChR was higher than that of ChR(P<0.01). Using trypan blue exclusion method, the results showed ADFMChR at various concentration of 1.0, 3.0, 10.0, 30.0, 100.0μmol/L had inhibitory growth effect to CoC1 cell,in a dose-dependent manner. ADFMChR at 30μmol/L could prolong CoC1 cell multiple time from 28.5h to 39.6h. The colony formation in soft agar assay showed that the inhibitory rate of colony formation was 12.6%,32.4%,56.6%,83.5%,90.1% and 75.3%,respectively after treatment with ADFMChR at various concentration of 1.0,3.0,10.0,30.0,100.0μmol/L and ChR at 100.0μmol/L, The formation inhibitory rate of ADFMChRwas higher than that of ChR(P<0.01). The ladder-shaped band could be shown in DNA agarose gel electrophoresis after treatment with ADFMChR at 30.0μmol/L for 48h and the ladder-shape band disappeared, which preincubated with GW9662 at 10μmol/L for 30 mins. Flow cytometry(FCM) analysis after PI stainning indicated that ADFMChR significantly induced apoptosis in a concentration- dependent, the rate of apoptosis was 33.1% and 73.7% respectively after treatment with 10.0, 30.0μmol/L of ADFMChR for 48h, which was higher than those at the same concentration ChR-treated cells(21.7%,40.0%)(P<0.01).Western-blot analysis indicated that exposure to ADFMChR at 0.3, 3.0, 30.0μmol/L for 24h,the protein level of PPARγand Bax in CoC1 cells was up-regulated by 10.9%,31.5%,48.5% and 4.8%,34.5%,65.1%; NF-κB and Bcl-2 in CoC1 cells was down-regulated by 5.0%,29.1%,43.5% and 10.5%,31.1%,56.5% respectively in comparision with the control group. After treatment with ADFMChR at 30.0μmol/L for 6,12 and 24h, the protein level of PPARγand Bax in CoC1 cells was up-regulated by 23.1%,54.0%,65.0% and 8.6%,27.9%,60.1% ; NF-κB and Bcl-2 in CoC1 cells was down-regulated 11.5%,29.0%,54.8% and 26.9%,49.4%,61.7% in comparision with the control group respectively.Conclusion:1) 5-allyl-7-gen-difluoromethyllenechrysin posses the inhibitory effect of the growth of CoC1 cells,in a dose-dependent manner.2) 5-allyl-7-gen-difluoromethyllenechrysin can significantly induce apoptosis of CoC1 cells.3) The induction of apoptosis of CoC1 cells by 5-allyl-7-gen- difluoromethyllene- chrysin may be mediated by activation of PPARγ, sequentially accompanied by reducing of protein levels of NF-κB and Bcl-2 and increasing of Bax expression.
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