Study On The Effects Of Estradiol And Progesterone On Growth Of Hela Cervical Adenocarcinoma Cell Lines In Vitro

[Objective] To investigate the effects of estradiol(E2) and progesterone(P) on growth of Hela human cervical adenocarcinoma cell lines in vitro and Preliminary to discuss their mechanism, so as to provide the basic theory of hormone replacement therapy(HRT) for the women of cervical adenocarcinoma after comprehensive treatment.[Methed] Hela human cervical adenocarcinoma cells were cultured in vitro and the experiment established the E2group, P group, E2combining with P group, and control group.(1)Used methyl thiazolyl tetrazolium (MTT) to detect the effects of E2, P and E2combining with P on proliferation of Hela cells in different concentrations(0.01μmol/L,0.1μmol/L,1μmol/L,10μmol/L,100μmol/L) and different time (24h,48h,72h) respectively, then obtaining the optimal concentration and time was1μmol/L and72h respectively, and determining the concentration of E2and P were all1μmol/L in E2combining with P group;(2)72h after acting with E2, P and E2combining with P, used inversion microscope, electron microscope and transmission electron microscope to observe the cell morphology of Hela cells;(3)72h after acting with E2, P and E2combining with P, used flow cytometry (FCM) to analyse the cell cycle, the ratio of apoptotic cell and the expression of Fas Ligand(FasL) protein.[Result](1)MTT showed①After24hours stimulation to Hela cells with different concentrations of E2, Hela cells was stimulated by10μmol/L of concentration(P<0.05), and there was no effect on Hela cells by other concentrations(0.01μmol/L,0.1μmol/L,1μmol/L,100μmol/L)(P>0.05); After48 hours stimulation to Hela cells with different concentrations of E2, Hela cells was stimulated by lμmol/L and lOμmol/L of concentrations(P<0.05), and there was no effect on Hela cells by other concentrations(0.01μmol/L,0.1μmol/L,l00μmol/L respectively)(P>0.05); After72hours stimulation to Hela cells with different concentrations of E2, Hela cells was stimulated by0.01μmol/L,0.1μmol/L,lμmol/L and10μmol/L of concentrations(P<0.05), but the stimulation didn't present concentration dependency, and there was no effect on Hela cells by concentration of100μmol/L(P>0/05); E2stimulated the growth of Hela cells and presented time dependency when the concentrations of E2was0.01μmol/L,0.1μmol/L,1μmol/L(P<0.05), that means with the extension of time, the stimulated effect increased;②P with all concentrations inhibited the growth of Hela cells significantly and presented time and concentration dependency(P<0.05), means with the extension of time or increase of concentration, the inhibition effect increased;③24h or48h with P combined with E2, when the concentration was100μmol/L, had apparent inhibition to Hela cells growth(P<0.05); There was no effect on Hela cells by other concentrations(0.01μmol/L,0.1μmol/L,1μmol/L,10μmol/L)(P>0.05);72h after P combing with E2, when the concentrations was10μmol/L and100μmol/L, had apparent inhibition to Hela cells growth(P<0.05); There was no effect on Hela cells by other concentrations(0.01μmol/L,0.1μmol/L,1μmol/L)(P>0.05);(2)Observed with inversion microscope, scanning electron microscope and transmission electron microscope we found that in the E2group, morphology of Hela cells was not significantly different compared with Control group; in the P group or P combined with E2group, the number and volume of cells were decreased, the speed of cell proliferation slowed down and apoptotic bodies appeared;(3)FCM showed in the E2group the phase of cell cycle was changed, that E2decreased the proportion of G0/G1phase(P<0.05), but there was no effect on the other phase of cell cycle or the apoptotic rate(P>0.05); In the P group or the P combined with E2group, the proportion of G0/G1phase and the apoptotic rate were increased and the proportion of S phase was reduced significantly, the role of P combining with E2was more remarkable than P(P<0.05);(4)Detecting of FasL with FCM showed FasL was expressed in all of groups, in the E2group, the expression of FasL-protein was no significantly different compared with Control group(P>0.05); In the P group or the P combined with E2group, the expression of FasL-protein was down-regulated significantly, and the role of P combining with E2was more remarkable than P(P<0.05).[Conclusion] E2stimulated the growth of Hela cells and presented concentration independency but time dependency, means with the extension of time, the stimulated effect increased. Progesterone inhibited the growth of Hela cells significantly and presented time and concentration dependency, means with the extension of time or increase of concentration, the inhibition effect increased. The lower concentrations of P combining with E2had no obvious stimulation or inhibition to the growth of Hela cells;and the higher concentrations groups showed inhibition to the growth of Hela cells, means that enough P can resist the role of proliferation enhanced by E2. In the P group or the P combing with E2group, the proportion of Go/G1phase and the apoptotic rate were increased significantly, the proportion of S phase was reduced. P induced apoptosis through blocking the phase of cell cycle. FasL-protein were high expressed in Hela cells. In P group or P combining with E2group, the expression of FasL-protein was down-regulated, means that P may against stimulating effect of E2through down-regulated the expression of FasL-protein.The role of P combining with E2was more remarkable than P,tip P can against stimulating effect of E2and induced apoptosis.