| Objective:In this experiment,ovarian cancer cells were cultured in vitro,and progesterone(P)and extracellular signal-rugulated kinase(ERK)inhibitor U0126 were used alone or in combination to treat different progesterone receptors.Progesterone receptor(PR)expression of ovarian cancer cell proliferation and exploration of Mitogen Activated Protein Kinase(MAPK)/ERK/Forkhead box O3A(FOXO3a)signaling pathway The molecular mechanism of action in ovarian cancer cells provides a theoretical basis for the treatment of ovarian cancer.Methods:1.Screening of ovarian cancer cell line HO-8910(high PR expression)and SKOV3(low expression of PR)as the research object,two ovarian cancer cell lines were grouped separately,and a blank control group was set up.The experimental group(P group,U0126 group,P)+U0126 group).The concentration of P group and U0126 group of two ovarian cancer cells was first defined.The concentration of P group was 0,1,10,20,40 ?mol/L,and the concentration of U0126 group was 0,10,20,40 ?mol/L,respectively.The drugs were used alone or in combination for 24,48,and 72 hours,and the proliferation of each group was detected by CCK8 method.2.Obtaining HO-8910 and SKOV3 ovarian cancer cells with good growth state,and interfering with the two cells with P(20?mol/L),U0126(10?mol/L),P(20?mol/L)+U0126(10?mol/L),respectively.After 24 hours of drug intervention,the growth state of each group was observed by inverted phase contrast microscope.Then,The cell proteins of each group were extracted and the expression of PR-A,PR-B,FOXO3 a,p-FOXO3 a,p-ERK proteins were detected by Western blot.Results:1.Different concentrations of P interfered with the survival rate of HO-8910 cells at different times: P promoted the proliferation of the two cells at a low concentration,inhibited the proliferation of the two cells at a high concentration,and the survival rate of the two cells appeared as the concentration of P increased and the duration of actionincreased.Time-dose-dependent reduction.The survival rates of the two groups of cells were compared: P at low concentration(1?mol/L)promoted the proliferation of ovarian cancer cells,and the proliferative effect on HO8910 cells was significantly higher than that of SKOV3(P<0.05);at high concentrations(10?20?40 ?mol/L),P inhibited the proliferation of ovarian cancer cells,and the effect of inhibiting proliferation of HO8910 cells was significantly higher than that of SKOV3(P<0.05).2.Different concentrations of U0126 interfered with HO-8910 cells for 24 h: U0126 could inhibit the proliferation of HO-8910 and SKOV3.SKOV3 showed a time-dose-dependent decrease in U0126.HO-8910 decreased the U0126 in a time-dose-dependent manner.When the concentration of U0126 was 10 and 20 ?mol/L,the proliferation activity did not change with time(P>0.05).3.P(20?mol/L),U0126(10?mol/L)combined intervention of ovarian cancer cells HO-8910,SKOV3 24 h after the survival rate decreased,the cell proliferation activity decreased compared with the intervention alone(P<0.05),and The effect of combined intervention on the inhibition of proliferation of HO-8910 cells was stronger than that of SKOV3 cells(P<0.05).4.P,U0126 alone and in combination with ovarian cancer cells HO-8910,SKOV3 for 24 h,Morphology of two ovarian cancer cells under inverted phase contrast microscope(magnification: 200×): control group HO-8910,SKOV3 cells showed epithelial adherent growth,showing out of the shape of round,fusiform,polygonal,paving stone arrangement,some cells have pseudopods;P,U0126 alone increased the cell gap,the number of adherent cells decreased,floating cells and cell debris increased,Interstitial cells can be seen.When P and U0126 were combined,the cell HO-8910,which was originally epithelial-like adherent growth,gradually became fusiform,and the border of SKOV3 cells gradually became blurred,and multiple cell fusion occurred.Under the above microscope changes,HO-8910 cells changed significantly compared with SKOV3 group.5.P(20?mol/L)and U0126(10?mol/L)alone and in combination with HO-8910 and SKOV3 cells for 24 h,the relative expression of FOXO3 a in the experimental group increased compared with the control group,and the expression of p-FOXO3 decreased in the control group(P<0.05);The increase of FOXO3 a in the combined intervention group was significantly higher than that in the drug alone group(P<0.05),and the decrease in p-FOXO3 and p-ERK was significantly lower than that in the drug alone group(P<0.05).In the same kind of cells,the relative expression of PR-A and PR-B protein in the experimental group was not significantly different from that in the control group(P>0.05).Conclusion:1.Progesterone may has a biphasic effect on the regulation of ovarian cancer HO-8910 and SKOV3 cells proliferation.Low concentration promoted ovarian cancer cell proliferation,and high concentration inhibited ovarian cancer cell proliferation;2.Progesterone may up-regulates FOXO3 a by inhibiting MAPK/ERK pathway,thereby affecting the proliferation of ovarian cancer HO-8910 and SKOV3 cells;3.The combination of progesterone and U0126 has synergistic effect on the inhibition of proliferation of ovarian cancer HO-8910 and SKOV3 cells;4.Progesterone inhibits the proliferation of ovarian cancer cells may be regulated by PR-B through the MAPK/ERK/FOXO3 a signaling pathway. |
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