Numerous physiological changes take place in a woman’s body during pregnancy. Throughout the long gestation period, the main energy pathway received by the fetus is glucose from the mother’s placenta (1). While pregnancy progresses, the fetal need for glucose grows, which leads to gestational diabetes mellitus (GDM) in women with otherwise healthy glucose metabolism or potentially impaired glucose tolerance prior to pregnancy (2, 3). As a common complication of pregnancy worldwide, uncontrolled GDM poses a serious threat to the mother, fetus, and newborn, increasing the likelihood of adverse pregnancy reactions (e.g., gestational hypertension, infection, and metabolic ketoacidosis) as well as malignant pregnancy outcomes (e.g., preterm abortion, neonatal hypoglycemia, and postpartum depression) (4). The focus of GDM prevention efforts is on preconception or early pregnancy, with only a minority of women with GDM requiring pharmacological treatment, and the identification of reliable underlying risk markers is valuable for the timely detection and prognosis of GDM (5).

The growth of a fetus from conception to successful delivery is a significant challenge for the physiological functions and regulatory systems of the mother (6). Patients with GDM frequently experience metabolic issues, such as increased insulin resistance, and generally suffer from systemic mild inflammation and immune dysregulation (7). In GDM, various types of immune cells, particularly regulatory T cells (Tregs), adapt spontaneously to prevent pregnancy interruption. Besides this, there is a proportional increase in circulating monocyte activation and an elevated level of cytokines, including IL-12 and IL-23, in mid to late gestation compared to non-pregnant women (8). It is quite predictable that changes in the quantity or function of immune mediators are involved in the development of GDM. Regrettably, the conclusions of the current studies on the correlation between maternal immune profiles and GDM are not entirely consistent, which may be due to factors such as differences in the samples and flaws in the design.

Nowadays, genome-wide association study (GWAS) and Mendelian randomization (MR) make it feasible to estimate the causal associations between immune traits and disease events on a large scale. GWAS identifies genome-wide sequence variation in specific human populations (9), whereas the existence of randomness in the process of genetic variation allows MR to be independent of common confounders and reverse causation (10). In this current study, by identifying single-nucleotide polymorphisms (SNPs) linked to complex traits, a comprehensive bivariate MR analysis was undertaken to identify causal relationships between immunocyte features and GDM. Figure 1 provides an illustration of this research.

GWAS studies have revealed associations between diseases and genetic variation, etc. Benefitting from a large, publicly available genomic data, our study attempted to offer genetic evidence for the causal link between GDM and multiple immune phenotypes. In this paper, we proved that four immune phenotypes have a significant causal association for GDM (P_FDR < 0.04), while GDM has the same relationship for five other immune phenotypes (P_FDR < 0.03).

As a globally prevalent obstetric disorder, GDM has been shown to be associated with many adverse maternal and fetal pregnancy outcomes. Abnormal maternal immune adaptation is key to the low-grade inflammation associated with the diagnosis of GDM, while immune cell infiltration of visceral adipose tissue causes the pathological dysregulation of insulin signaling and contributes to insulin resistance. Specifically, CD39 is located on the surface of trophoblast cells in the normal human placenta and regulates ATP-dependent trophoblast function, which is critical for immune tolerance and the maintenance of a normal pregnancy (24). Several studies have shown that measuring CD19+ subpopulations can help predict the pregnancy outcome in women, with a trend towards lower peripheral CD19+ B cells in women who miscarry compared to those who subsequently give birth (25). The frequency of CD19(+) CD5(+) cells was also significantly increased in the peripheral blood of patients with pre-eclampsia compared to normal pregnant women (26). In addition, the maternal humoral response to fetal anti-HLA-DR immunoglobulin antibodies may influence the development of pregnancy-induced hypertension (27).

Our results demonstrate that the elevated levels of HLA DR on plasmacytoid DC and HLA DR++ monocyte %leukocyte increase the risk of GDM. As a specialized antigen-presenting cell, dendritic cells (DCs) regulate the immune response and bridge the gap between innate and adaptive immunity (28). DCs play crucial roles in the growth and development of embryos and fetuses in the mother’s womb; dysregulation of the DC subpopulations appears to be linked to adverse pregnancy outcomes (29). HLA-DR is the most common MHC class II molecule on the surface of antigen-presenting cells. Specially, the expression of HLA-DR on the surface of DCs increases the abundance of protein complexes and is accompanied by the production of co-stimulatory molecules and cytokines (30). Moreover, recent studies have demonstrated that the level of HLA-DR is an indicator of monocyte immunocompetence, which not only assists in antigen presentation but also strengthens TLR-2-mediated signaling, cell proliferation, and maturation (31).

On the contrary, the growing levels of CD127 on CD28+ CD45RA+ CD8br and CD19 on PB/PC decrease the risk of GDM. The reduced activity of inhibitory Treg isoforms in GDM was pointed out to be associated with the upregulation of pro-inflammatory factor concentrations which include IL-6 and TNF-alpha (32). Another study showed that the percentage of circulating Treg subpopulation cells defined by CD3+CD4+CD25 bright/dim CD127 expression was reduced in GDM pregnancies compared with glucose-tolerant pregnancies (33). CD19 is a transmembrane protein on the surface of B cells, which is tightly connected with B cell activation, signaling, and growth regulation. For IgG4-related diseases, the peripheral blood was significantly enriched in B cell populations, including CD19+ CD24-CD38hi PB/PC. After glucocorticoid administration, the levels of these cells declined, accompanied by an improvement in clinical symptoms (34). In summary, our research indicates that immune cells have a significant role in GDM’s early diagnosis, therapeutic monitoring, disease activity assessment, and adaptive therapies.

On the other hand, we also revealed that the percentage of various immunological phenotypes was altered as a result of GDM. Strikingly, we found that GDM commonly lowered the abundance of CD39+ activated Treg %CD4 Treg, CD39+ activated Treg AC, CD39+ resting Treg % CD4 Treg, CD39+ resting Treg AC, and CD39+ CD8BR %T cell. Extracellular ATP is an effective proinflammatory factor in vivo, and its hydrolysis is important for its immunosuppressive function. As an extracellular ectonucleotidase, CD39 has been implicated as a major marker of FOXP3+ Treg and cleaves ATP to form AMP in the rate-limiting step. It is notable that the percentage of D39+ Treg cells was significantly decreased in type 2 diabetes patients as compared to the controls. In vivo experiments have shown that CD39-deficient mice exhibit impaired glucose tolerance in an oral glucose tolerance test (35). The supplementation of soluble CD39 to pre-diabetic NOD mice reduces the extent of extracellular ATP, inhibits the multiplication of CD4+ T cells, and delays the further progression of diabetes (36). It reminds us that individualized treatment for CD39 is probably a promising option for pregnant women.

Apparently, our study offers a foundation for delineating the intricate causal association between immune cells and GDM. However, there are still several limitations in our work. Firstly, although we performed MR analyses with a large-scale GWAS cohort and avoided potential confounders or reverse causation, genetic heterogeneity among different human populations still attenuates the credibility and validity of the GWAS results. Secondly, when examining the association of immune cells with GDM, a more relaxed threshold was chosen to ensure accurate data on SNPs. Even with the FDR test applied, this may still lead to a minor bias in the results. Finally, for GDM, our study was unable to further probe specific traits (for example—age, weight, and hormone levels) in the group of pregnant women.

In summary, we emphasized the causal relationship between a number of immune phenotypes and GDM through a full bidirectional MR analysis. To our knowledge, this is the first MR analysis carried out between immune phenotypes and GDM, providing novel insights into understanding the delicate balance between maternal immune mediators and GDM. GDM is a complicated and dynamic condition, and the pathophysiological mechanisms are not fully clarified. This research enables researchers to better explain the physiological mechanisms, with a view to filtering and monitoring high-risk groups for GDM, contributing to the early intervention and the development of new treatments of GDM.

The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/ Supplementary Material .

ZJ: Conceptualization, Data curation, Investigation, Visualization, Writing – original draft. CZ: Conceptualization, Data curation, Visualization, Writing – original draft. JY: Supervision, Validation, Writing – review & editing. QH: Validation, Visualization, Writing – review & editing. XZ: Validation, Visualization, Writing – review & editing. DY: Validation, Visualization, Writing – review & editing. NX: Formal Analysis, Funding acquisition, Project administration, Supervision, Writing – review & editing. JC: Formal Analysis, Funding acquisition, Project administration, Supervision, Writing – review & editing.