Pregnancy is a complex physiological process, all the subjects included in this study were pregnant. Previous studies mostly compared the pregnant women with URSA history and normal gestational age, while the pregnant women with URSA and normal gestational age were compared in this study. The two groups of people had a common immune basis and were all pregnant. An identical immune basis between each group is a necessary condition for effective experimental results, and observation of the expression status of Th1/Th2 during pregnancy can more truly and effectively reflect the immune status and relative number of Th cells in patients with unexplained recurrent abortion.
Studies[20] have shown that the first three months of pregnancy is closely related to inflammation and may produce corresponding immune rejection. In the second trimester, anti-inflammatory and Th2 immune microenvironment are the main factors, which are necessary for fetal growth. There is a transition between Th1 and Th2 immune states between the first and second trimester of pregnancy, which is a necessary transition process for successful delivery. However, patients with URSA may show a continuous inflammatory immune response in the second trimester of pregnancy, which cannot be adjusted by increasing inhibitory immune mechanism, eventually leading to the intrauterine microenvironment with persistent inflammation, which is harmful to the fetus and leads to abortion.
various types of immune cells may be involved in promoting immune tolerance during pregnancy, the immune cells in the decidual tissues of the mother produce th2-related cytokines, including il-3, il-4, il-5, il-6, il-9, il-10 and il-13 [21],which can promote the growth of trophoblast cells and may be conducive to maintaining the success of pregnancy[22-25], and some studies suggest that Th1-type inflammatory response may lead to embryo loss[26, 27],Th1 cells are activated and release cytokines, including IFN-γ, TNF-α and IL-2,and these cytokines mainly mediated immune cells and local inflammatory response[28].
In addition to setting the Pregnancy group as the control group, the ad-pregnancy group was added as the other control group in this study. The recruiters in the ad-pregnancy group had a history of spontaneous abortion, stillbirth, or stillbirth, and B-mode ultrasound indicated gestational status, but excluded chromosomal abnormalities, intrauterine infections, autoimmune diseases, endocrine/metabolic disorders, thrombo-hemostatic protein abnormalities, and uterine abnormalities. With insufficient number of embryo losses to meet the criteria for diagnosis of URSA, we might consider the ad-pregnancy group to be the advance-ursa group. Further validation of the ad-pregnancy group with the URSA group, the pregnancy group. Our experimental research results showed that the ratio of il-2 / il-4 and IFN-γ/il-4 in peripheral blood of URSA group was significantly higher than that of Pregnancy group(P<0.05), the ratio of IFN-γ/il-10 and IFN-γ/il-4 in peripheral blood of ad-pregnancy group was significantly higher than that of pregnancy group, it is consistent with the results of other people's research[11, 21, 29-31], these results suggested that th1 cytokines increased and th2 cytokines decreased in the URSA patients.
The results of this paper indicated that, with the exception of il-6, there was no statistically significant difference between the URSA group and the ab-pregnancy group, the results of our study showed that, when compared with the URSA group, the peripheral blood of the ad-pregnancy group had a higher il-6 content and had a statistically significant difference.
When we looked at the ab-pregnancy group as the advance-ursa group, this result was not in line with expectations, that was probably because il-6 is a nonspecific infection markers, normal serum il - 6 in the body content is relatively low, when some diseases occur, the content of il - 6 will rise sharply in the body, especially when the body infection, and at the time of the recruiting research object of this study, not to conduct a comprehensive physical examination, there may be some potential infection or other diseases affect the content of il - 6 in the body , perhaps the increasing of il-6 in ad-pregnancy group may be due to individual inconsistencies in the exclusivity response to foreign antigens.
The results of this study showed that when compared with the pregnancy group, the peripheral blood of the ab-pregnancy group had a higher il-6 content and had a statistically significant difference and from the level of il-6 in peripheral blood of the three groups (Fig. 2-c), it can be seen that the content of URSA group and ab-pregnancy group is higher than that of pregnancy group, the results of Lim KJ's study[32] showed that the level of IL-6 in the decidual tissue of URSA patients was low, which was inconsistent with the results of this study, possibly because IL-6 is a proinflammatory factor, which can promote the maturation and differentiation of T and B lymphocytes, and promote the immune activity of NK cells and CTL cells.IL-6 further promotes the production of other cytokines and inflammatory mediators, and interacts with each other, so that the balance between inflammatory factors and anti-inflammatory factors is out of control and the inflammatory reaction is excessive, and the accumulation of neutrophils in the inflammatory site leads to more inflammatory mediators, which further damages trophoblast cells and is not conducive to embryonic development.
But in some experiments[33] , it was found that the content of IL-6 and mRNA in the decidual tissues of URSA patients increased, the results are consistent with the results of our study. However, il-6 is a Th2 type cytokine, these results contradicted the th1/th2 equilibrium theory of maternal-fetal interface.
Previously, it was thought that the etiology of multiple miscarriages in URSA patients is that immune response from the Th2 immune response as the main guide to the Th1 immune response as the leading metastasis[34].However, the results of our study showed that the reason of embryo loss was not only the transfer of immune response, but also the inflammation of maternal-fetal interface.
The results of our study showed that there was no significant difference between the groups of il-10 and TNF-α, however, there were studies[35-38] that showed that the expression level of TNF-α in URSA patients increased as the number of miscarriages increased, indicating that the increase in cytokines was positively correlated with the increase in the number of miscarriages and TNF-α might be an important reference for the results of pregnancy, and the data of Marzi M study suggested that the production of il-10 in peripheral blood monouclear cells decreased during spontaneous abortion, manifesting that il-10 may be a protective factor in pregnancy[27]. In this paper, we can't get similar results in our data, this may be because the number of samples included in this study is slightly small, or it may be because there are differences in environmental genes between different countries, ethnic groups or regions. Studies have[39-41] proved that unexplained recurrent abortion may be related to gene polymorphism.
ROC curve was plotted from data of URSA group and Pregnancy group in this study. The AUC of IFN-γ/il-4 was 0.821, with high diagnostic efficiency, sensitivity as high as 84.09% and specificity as high as 69.05%, which were higher than other indicators, the initial diagnosis of URSA with this index could reduce the rate of missed diagnosis.
However, it was found in the study that there was no statistical difference in the content of il-4, il-2 and IFN-γ in peripheral blood between each group, but the ratio of IFN-γ/il-4 and il-2/il-4 in peripheral blood of URSA group was significantly higher than that of Pregnancy group. This may be because the decreased or increased amount of il-2, il-4 and IFN-γ in patients is not statistically significant compared with Pregnancy group, but the ratio of the two variables is equal to the combination of the differences between patients and normal pregnant women, so that the new variable has obvious differences between the two; cytokines may be because of the complexity of the interaction of the formation of the complex network of cytokines to adjust a single factor can be the rest is influenced by many kinds of cell factors or other factors, when the two effects opposite factors taken together, can eliminate some indirect interactions between cytokines, amplifying the differences between different groups; it may also be because the sample size included in this study is slightly smaller.