Systemic immune differences in patients undergoing frozen embryo transfers that result in live birth

To determine if there are systemic differences in cytokines and angiogenic factors on the day of frozen euploid embryo transfer (FET) between cycles that result in live birth versus those that do not. 20 patients undergoing FET of euploid embryos were enrolled in the study. Blood samples were collected on the day of FET and 9 days later. If pregnancy was confirmed on day 9, additional blood samples on day 11 post FET and just prior to the first trimester ultrasound were collected. Plasma and peripheral blood mononuclear cells (PBMCs) were isolated for analysis. Plasma levels of 60 cytokines and angiogenic factors were measured using a multiplex bead-based assay, and real-time PCR was used to measure the expression of 92 immune-related genes in PBMCs. Protein concentration and gene expression were compared between the cycles that resulted in a live birth and those that did not using Mann-Whitney test, and comparisons over time were assessed using repeated measures ANOVA. The 20 participants underwent a total of 22 frozen euploid embryo transfer cycles, and 11 cycles resulted in a live birth. Of the 60 plasma proteins evaluated on the day of FET, two were significantly lower in cycles that resulted in a live birth compared to those that did not: Interleukin (IL)-13 (106.4 ng/mL vs. 207.5 ng/mL, p=0.047) and macrophage derived chemokine (MDC; 344.9 ng/mL vs. 501.1 ng/mL, p=0.013). When IL-13 and MDC levels were examined in PBMCs collected at all time points in early pregnancy, their levels remained unchanged across all time points. Three of 92 immune related genes were over-expressed in PBMCs from the day of FET in cycles that resulted in a live birth versus those that did not: CCR5 (median 1.83 vs 1.02, p=0.026), CD8A (median 1.46 vs 1.08, p=0.041), and SMAD3 (median 1.33 vs 1.01, p=0.015). This study provides a broad assessment of the systemic immune status of patients undergoing FET. While we observed gene and protein changes in immune cells and markers between cycles that resulted in a live birth compared to those that did not, many measured targets were unchanged. This suggests that the local immune milieu may be more significant for implantation and pregnancy outcomes than what is altered systemically.