| Objective:The conditions and action mechanism of growth hormone promoting the proliferation of endometrial cells were explored at the cell level,and then the thin endometrial rat model was established.While exploring the proliferation effects of growth hormone promoting the proliferation of endometrial cells and the proliferation promotion mechanism of growth hormone at the animal level,vitamin E was innovatively added.Through experimental means such as morphology and molecular biology,the therapeutic effects of vitamin E and its combination with growth hormone on the model rats were observed and recorded,and the proliferation promotion mechanism of vitamin E was also explored.Methods:1.Cytological experiment: The endometrial cells were cultured with GH at different time and concentration gradients in vitro to explore the conditions of GH promoting the proliferation of endometrial cells.RT-PCR and Western Blot experiments were used to detect the expressions of key proteins and transcription factors of Wnt signaling pathway,to verify the correctness of the exploration culture conditions and to explore the mechanism of GH promoting the proliferation of endometrial cells at the cell level.2.Morphological experiment: A total of 90 rats with normal estrous cycles were randomly divided into the control group,corn oil group,model group,GH group,Vit.E group,and the GH and Vit.E combined treatment group.The rats in the control group were raised conventionally without treatment.The rats in the corn oil group were not treated and were given corn oil gavage for 14 days on the next day of estrous.The TE models were established in the model group,GH group,Vit.E group and the combination treatment group by intrauterine perfusion with 95% ethanol on the day of estrous.Normal saline was injected into uterine cavity once after operation in model group.In the GH group,intrauterine infusion of GH was performed once on the day after surgery.Vit.E was given intragastrically for 14 days on the next day in the Vit.E group.The rats in the combination treatment group were given intrauterine infusion of GH once on the day after surgery and received Vit.E continuous gavage on the next day for 14 days.After 14 days,they were sacrificed in estrous.The uterine tissue sections were prepared.HE staining was used to directly observe the thickness of endometrium,the number of glands,and the number of blood vessels in rats,to confirm the success of TE model establishment and to observe the therapeutic effects of GH and Vit.E on model rats.3.Western Blot: All the proteins in uterine tissues of rats were extracted,and the expression levels of landmark proteins in endometrial epithelial cells and stromal cells were detected to indirectly judge the condition of endometrium.The success of TE model establishment was detected at the molecular level,and the therapeutic effects of GH and Vit.E on model rats were observed.At the same time,the key proteins and transcription factors in the Wnt signaling pathway were detected to explore the mechanism of GH and Vit.E in promoting the proliferation of endometrial cells.Results:1.Cell experiments showed that the viability of epithelial cells and stromal cells was increased when GH was 200 pg/m L for 48 h.However,the uterine epithelial cells cultured at800 pg/m L GH concentration for 48 h in vitro had the best proliferation-promoting effect,and the stromal cells cultured at 800 pg/m L GH concentration for 24 h in vitro had the best proliferation-promoting effect.After GH stimulation,the expression levels of key proteins and transcription factors of Wnt signaling pathway in endometrial cells were significantly increased(P < 0.05).2.The endometrial thickness,the number of glands and the number of blood vessels in the model group after intrauterine infusion with 95% ethanol were significantly lower than those in the control group(P<0.05).3.There was no significant difference in uterine cavity space between the control group and the corn oil group,and the corn oil did not interfere with the therapeutic effect of Vit.E group.The endometrial thickness in the model group was the lowest.The endometrial thickness in the GH group was significantly higher than that in the model group,but slightly lower than that in the control group.The endometrial thickness in the Vit.E group and the combination treatment group was significantly higher than that in the control group,and the endometrial thickness in the combination treatment group was the highest(P<0.05).Gland number in the model group was significantly lower than those in the control group,Vit.E group and the combination treatment group,while those in the GH group and Vit.E group were slightly lower than those in the control group(P<0.05),with no significant differences among the other groups(P>0.05).For the number of endometrial blood vessels,the number of endometrial blood vessels in the model group was significantly lower than that in the control group and the combination treatment group,and there was no significant difference between the other groups(P>0.05).4.Western Blot results showed that the expression levels of landmark cytokeratin and vimentin in endometrial epithelial cells and stromal cells of rats in the model group were the lowest and significantly lower than those in the other groups.The protein expression of GH group was slightly lower than that of the control group.The protein expression level in the control group was slightly lower than that in the Vit.E group and the combination treatment group,and the protein expression level in the combination treatment group was the highest(P<0.05).The expressions of β-catenin,transcription factors MET and c-MYC,which are key proteins in Wnt signaling pathway,were detected.The results showed that the expressions ofβ-catenin,MET and c-MYC in model group were significantly lower than those in GH group and Vit.E group(P<0.05).Conclusion:1.The cell viability was significantly increased when the endometrial epithelial cells and stromal cells were cultured in vitro for 48 h at GH concentration of 200pg/m L.However,the endometrial epithelial cells cultured for 48 h with an additional GH concentration of 800 pg/m L had the highest survival rate,and the endometrial stromal cells cultured for 24 h with an additional GH concentration of 800 pg/m L had the highest survival rate.2.Simple GH intrauterine infusion and Vit.E gavage can significantly increase the endometrial thickness of TE rats,and the combination of GH intrauterine infusion and Vit.E gavage has better therapeutic effects.3.GH and Vit.E can regulate the Wnt/β-catenin signaling pathway and promote cell proliferation by affecting the expression of transcription factors MET and c-MYC. |
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