| Objective: Studies have found that menstrual blood-derived mesenchymal stem cells can effectively repair endometrial damage,however its mechanism is still unclear.In this paper,the changes of Interferon-gamma、Interleukin-2、Interleukin-6 in the process of repairing damaged endometrium by MenSCs would be discussed,which would further explore its immunomodulatory mechanism.Methods:1.To establish the mouse model for damaged endometrium.The vaginas of mice were washed with normal saline to take the secretions at the same time every day.The vaginal secretions of mice were detected by HE staining method to determine the stage in estrous cycle.The model of damaged endometrium was established by intrauterine infusion of 95% ethanol for 15 s in the afternoon on the day of estrus.After three estrous cycles,uterine tissue was collected in the afternoon on the day of estrus,and the morphology and thickness of endometrial tissue were observed by HE staining.2.To study the immunomodulatory effects of Th 1/Th 2 cells by MenSCs in repairing damaged endometrium.The experiment was divided into three groups: normal control group,PBS transplantation group and MenSCs transplantation group.Normal control group: no damage in the endometrium,PBS transplantation group: PBS was transplanted in situ into the damaged endometrium model immediately after intrauterine infusion of 95% ethanol for 15 s,MenSCs transplantation group: MenSCs was transplanted in situ into the damaged endometrium model immediately after intrauterine infusion of95% ethanol for 15 s,and the uterine tissues were collected after three estrous cycles after transplantation.The expressions of IFN-γ,IL-2 and IL-6 in the endometrium of the three groups were measured by Western Blot and RT-q PCR method.This led to knowledge MenSCs’ Immunomodulatory mechanism for repairing damaged endometrium.Results: 1.Detection the stage of estrous cycle in mice: In proestrus,a large number of nucleated epithelial cells and a small number of keratinocytes were found in vaginal secretion smear,during estrus,keratinocytes and a few nucleated epithelial cells could be seen in the whole field in vaginal secretion smear,in metaestrus,a small amount of keratinocytes and leukocytes could be seen in vaginal secretion smear,During anestrus,a lot of leukocytes could be seen in vaginal secretion smear.2.HE staining results: in the damaged endometrium group,the endometrium was significantly thinner,the luminal epithelium of endometrium was damaged or even broken,and the number of glands was reduced,which proved that the model was successfully established.3.Western blot was used to detect the protein expression of immune factors.The expression of IFN-γ in the MenSCs transplantation group was lower than that in the PBS transplantation group(P<0.05),and there was no significant difference between the MenSCs transplantation group and the normal group,while the expression of IFN-γ in the PBS transplantation group was higher than that in the normal control group(P<0.05).The expression of IL-2 in MenSCs transplantation group was lower than that in PBS transplantation group(P<0.01)and higher than that in normal control group(P<0.01),while IL-2 in PBS transplantation group was higher than that in normal control group(P<0.01).The expression of IL-6 in the MenSCs transplantation group was higher than that in the PBS transplantation group(P<0.05),but there was no significant difference between the MenSCs transplantation group and the normal group.The expression of IL-6 in the PBS transplantation group was lower than that in the normal control group(P<0.01).4.The m RNA expression of immune factors was detected by PCR.The m RNA expression of IFN-γ in the MenSCs transplantation group was lower than that in the PBS transplantation group(P<0.05),but there was no significant difference between the MenSCs transplantation group and the normal group,while the m RNA expression of IFN-γ in the PBS transplantation group was higher than that in the normal control group(P<0.05).The m RNA expression of IL-2 in MenSCs transplantation group was lower than that in PBS transplantation group(P<0.01)and higher than that in normal control group(P<0.01),while IL-2 in PBS transplantation group was higher than that in normal control group(P<0.001).The m RNA expression of IL-6 in the MenSCs transplantation group was higher than that in the PBS transplantation group(P<0.05),but there was no significant difference between the MenSCs transplantation group and the normal group.The m RNA expression of IL-6 in the PBS transplantation group was lower than that in the normal control group(P<0.05).Conclusions:1.After damage with 95% ethanol,the expression of pro-inflammatory cytokines IFN-γ and IL-2 become higher in mice,showing that Th 1 cell is the main inflammatory reaction,and it may inhibit the anti-inflammatory factor IL-6 which protects the immune tolerance of the body.2.The mice after MenSCs transplantation represent higher expression of IL-6,showing that Th 2 cell is the main anti-inflammatory reaction,which is beneficial to endometrial repair,and may also inhibit the abnormal immune reaction induced by IFN-γ and IL-2.3.The endometrial inflammation still exists after MenSCs transplation,so the MenSCs transplantation therapy should be further improved. |
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