Express Of Akt/GSK3? Signal Pathway In The Process Of Endometrial Disintegration And Repair In Mice Menstrual-like Model And Its Regulation By Progesterone

Objective: To explored the role of this pathway in the process of menstrual endometrial disintegration and repair.To detect the expression level of protein kinase B(Akt)and glycogen synthase kinase 3 beta(GSK3?)in the uterine horn tissue in the process of menstrual-like changes in mouse model by physiological progesterone withdrawal,and the effect of progesterone replacement on ATK/GSK3? signaling pathway before and after the critical period of menstruation(12 h and 16 h after progesterone withdrawal).Methods: 120 health adult ovariectomized C57BL/6J female mice were treated with estrogen and progesterone to mimic physiological hormone fluctuations.Oil was injected into the uterine luminal to induce endometrial decidualization.P4 explant was removed after 49 h to induce mouse menstrual-like changes and this point was defined as 0 h.Ninety mice were randomly divided into nine groups,which were sacrificed by cervical dislocation at 0 h,8 h,12 h,16 h,20 h,24 h,32 h,40 h,and 48 h after progesterone withdrawal(The physiological progesterone withdrawal cycle in mice is 48 hour)to collect uterine horns.The lefted thirty mice were randomly divided into three groups,1216 h group(progesterone withdrawal for 12 h and progesterone replacement for 16 h),1224 h group(progesterone withdrawal for 12 h and progesterone replacement for 24 h),and 1624 h group(progesterone withdrawal for 16 h and progesterone replacement for 24 h).After sacrificed,the uterus horns were collected to detect both the m RNA and protein levels of Akt and GSK3? m RNA by Real-time PCR,Western Blotting and immunohistochemistry.Results: 1.Express of Akt/GSK3? signal pathway in the process of endometrial disintegration and repair in mouse menstrual-like model.No obvious differences were observed in all groups about expression levels of total Akt proteins in endometrial tissues(P>0.05).After progesterone withdrawal,the expression levels of Akt m RNA and p-Akt(Ser473)proteins were gradually decreased and then gradually increased after reaching the minimum at 20 h.On the contrary,the expressions of GSK3? m RNA and p-GSK3?(Ser9)protein levels were increased and reached a peak at 8 h(P<0.05)before underwent a gradually declined process(8 h~24 h)followed by an increased process(24 h~48 h).Immunization showed that p-Akt(Ser473)and p-GSK3?(Ser9)proteins in mice uterine tissues exhibited similar distribution and were mainly expressed in the stroma cell,glandular epithelium cell and luminal epithelium cell,and concentrated distributed around the decidualized region.2.The effect of rogesterone replacement in the early stage of menstruation on the ATK/GSK3? signaling pathway.The epression of Akt and GSK3? were significant corelated in three groups of early stages of menstruation(12 h group,1216 h group and 16 h group)(r=0.819,p<0.01).The expression level of Akt and GSK3? of 1216 h group were approximately equal to those of 16 h group(p>0.05),but lower than those in 12 h group(P<0.05).Total Akt proteins in uterine tissues of different groups were not significant differente.The expression trends of p-Akt and p-GSK3? were consistent(r=0.867,p<0.01).Compared with the 12 h group,the expression of p-Akt and p-GSK3? protein was significantly higher in 1216 h group(p<0.05).But the expression of both proteins was lowest in 16 h group.3.The effect of progesterone replacement on Akt/GSK3? signaling pathway in the late stage of menstruation.The m RNA levels of Akt and GSK3? and the protein level of total Akt exhibited on significant difference among three groups of late stage of menstruation(1224 h group,1624 h group and 24 h group).The protein of P-Akt and p-GSK3? showed stronge expression in 1224 h group,but lower in 1624 h group and barely detected in 24 h group.Conclusion: 1.Akt/GSK3? signal pathway maybe play a crucial role in endometrial disintegration and the following recovery process in mouse menstrual-like model.2.Reverses menstruation by rapid activating the Akt/ GSK3? signaling pathway and its downstream cascade reaction through non transcriptional mechanisms when progesterone was added back at 12 h after withdrawal,but at 16 h could not be reversed completely.