ERα Inhibits Uterine Smooth Muscle Cell Differentiation Induced By Myocardin

Objective:1. Study the expression of the differentiation marker gene induced by myocardin inuterine smooth muscle cell.2. Investigate ERα-induced proliferation of uterine smooth muscle cell.3.Study ERα-inhibited transcription activation and expression of the differentiationmarker gene which are induced by myocardin in uterine smooth muscle cell.4. Study the protein-protein interaction between ERα and myocardin.Methods:1. Establishing the SD rat model of unterine leiomyoma.2. Determining the expressions of myocardin, SM22α, ERα, PCNA in uterinesmooth muscle tissues of leiomyoma and normal SD rats by IHC, RT-PCR, westernblotting.3.In primary cell culture hysteromyoma and normal uterus smooth muscle， theactivity of SM22-LUC、SM22mut-LUC、SM22Emut-LUC and the expression of thesmooth muscle differentiation and proliferative marker gene are detected.4.In COS-7cell culture, the interaction of myocardin and ERα was detected byco-IP. The expression of myocardin and ERα was detected by Immunofluorescence.After ERα was transfected in normal smooth muscle cells，the protein and mRNAexpression of myocardin were detected by RT-PCR and western bloting.Results:1. The model of hysteromyoma are verified by the gross appearance and the HEstaining.2.RT-PCR、 western bloting、 IHC suggested that the expressions of both myocardin and the differentiation marker SM22α were higher in the normal uterussmooth muscle tissue than in the hysteromyoma tissues.On the contrary，the expressionsof both ERα and the proliferation marker PCNA were higher in hysteromyoma tissuethan the normal uterus smooth muscle tissue.3.In the primary cell cultures,myocardin alone is able to strongly drive theactivities of the SM22promotesr and SM22Emut promoter，but have no function forSM22mut promotr. But ERα inhibited the luciferase activity which was driven bymyocardin.4. The direct interaction between ER and myocardin was verified by Co-IP. Theinhibitory effect is correlated with the concentrations of the ERα.Conclusions:1. Myocardin activates transcription and expression of differentiation markerSM22α through binding with CArG box in USMCs.2. ERα inhibited USMCs differentiation through downregulating myocardin-inducedtranscription and SM22α expression.3. The interaction between myocardin and ER may facilitates the development ofunterine leiomyoma.