Correlation Studies Between Vitamin D Signal At Human Materno-Fetal Interface And Recurrent Spontaneous Abortion

PART ?The expression of vitamin D pathway related factors in materno-fetalinterface of recurrent spontaneous abortion patients and its relationshipwith IL-23/IL-17 inflammation axisObjective Analyze the relationship between vitamin D pathway related factors such as 25(OH)D,VDR and CYP27B1 and IL-23/IL-17 inflammation axis related factors and recurrent spontaneous abortion in the first trimester.Methods Patients with recurrent spontaneous abortion and underwent artificial abortion early pregnancy were recruited.The expressiones of VDR and CYP27B1 protein were detected by Western blotting.The levels of 25(OH)D,IL-23,IL-17,TGF-beta in decidual tissue were measured by Elisa.Finally,the correlation of with IL-23/IL-17 inflammation axis factors,and the relationship between 25(OH)D and the risk of recurrent spontaneous abortion were also analyzed.Results(1)The expression of VDR and CYP27B1 protein were detected in all the decidua tissues of the two groups.(2)Compared with the normal early pregnancy group,the expression of VDR in the decidua tissue of the RSA group was significantly decreased(P<0.05),but the difference between CYP27B1 between the two groups was not statistically significant(P>0.05).(3)Compared with the normal pregnancy group,the levels of 25(OH)D and TGFbeta in the RSA group were decreased,and the levels of IL-23 and IL-17 were increased,and both the difference was statistically significant(P<0.05).(4)Correlation analysis showed that 25(OH)D had a significant negative correlation with IL-23,and IL-17 was positively correlated with IL-23(P<0.05).(5)logistic regression analysis showed that 25(OH)D was positively correlated with the risk of recurrent spontaneous abortion(P<0.05).Conclusion There is abnormal vitamin D pathway in maternal-fetal interface in recurrent spontaneous abortion.Vitamin D pathway is correlated with IL-23/IL-17 inflammatory axis related factors and the pathology of recurrent spontaneous abortion.PART IIThe effect of 1,25-(OH)2D3 on IL-23/IL-17 axis related factors in humandecidual immune cells of the first trimesterObjective Analyze the effects of 1,25-(OH)2D3 on the IL-23/IL-17 axis related factors in human decidual immune cells(DICs)of the first trimester.Methods The decidua tissues of patients with RSA and normal pregnancy were collected,and decidual cells were isolated and for primary cell culture.The DICs were stimulated by solvent(ethol)and physiological concentration of1,25-(OH)2D3(10-7M)for 24 hours,respectively.Then,the supernatant of cell and cell culture was collected.The percentage of Th17 in each group was analyzed by flow cytometry.The expression of Foxp3 and ROR gamma t m RNA in each group was detected by real-time PCR.The levels of IL-23,IL-17,TGF-beta in the supernatant of each group was detected by Elisa.Results(1)Compared with the control group,the proportion of Th17 is increased,the expression of ROR gamma t m RNA increased,the expression of Foxp3 m RNA decreased,and the levels of IL-23 and IL-17 increased in RSA group,and all the differences were statistically significant(P<0.05).(2)1,25-(OH)2D3 can reduce the proportion of Th17?the level of IL-23 and IL-17?the expression of ROR gamma t m RNA,but improve the expression of Foxp3 m RNA,all the differences were statistically significant(P<0.05).(3)The leveles of TGF-beta in the supernatant of cell culture were lower than those of the sensitivity test of the kit,and could not be detected.Conclusion 1,25-(OH)2D3 may play a protective role in pregnancy by inhibiting the related factors of the in the decidua immune cells.PART III The effect of 1,25-(OH)2D3 on human decidual stromal cells(DSCs)of thefirst trimesterObjective To analyzhe the effects of 1,25-(OH)2D3 on IL-23/IL-17 axis related factors?MMP-9/TIMP-3 and HOXA-10 in DSCs of the first trimester.Methods The DSCs were isolated from the decidua tissue of the patients with RSA and normal early pregnancy.After be identified by antecedent immunocytochemistry,the DSCs were treated with solvent(ethol)and physiological concentration 1,25-(OH)2D3(10-7M)for 24 hours respectively.Then,the cells and cell culture supernatant were collected.The expressions of TIMP-3 and HOXA-10 m RNA in each group were detected by real-time PCR,and the levels of IL-23,IL-17 and MMP-9 in cell culture supernatants were detected by Elisa.Results(1)It was identified that the purity of the DSCs was close to100%,which was in line with the experimental requirements.(2)Compared with the control group,the expression of HOXA-10 m RNA in DSCs decreased in RSA group,and the level of IL-17 and MMP-9 in the supernatant increased(P<0.05).The difference was statistically significant,but there was no significant difference in the expression of TIMP-3 m RNA between the two groups(P>0.05).(3)1,25-(OH)2D3 can reduce the secretion of IL-17 and MMP-9 and improve the expression of HOXA-10 m RNA and TIMP-3 m RNA significantly(P<0.05).(4)The leveles of IL-23 in the supernatant of cell culture were lower than those of the sensitivity test of the kit,and could not be detected.Conclusion 1,25-(OH)2D3 may play a protective role in pregnancy by inhibiting the secretion of IL-17 and MMP-9 while improving the m RNA expression of HOXA-10 and TIMP-3 from the decidual stromal cells.