Measuring Immune Tolerance to Predict Miscarriage or Failed Embryo Transfer

There is increasing evidence of the importance of T cell immunoglobulin and mucin-containing protein 3 (Tim-3) in suppressing allograft rejection, and thus it is hypothesized to play a role in pregnancy. Galectin-9 (Gal-9) is a ligand for Tim-3 activation which promotes Th1 apoptosis signaling. Activation of Tim-3 by Gal-9 has also been shown to suppress NK cell cytotoxicity at the maternal-fetal interface. Previous studies have evaluated Tim-3 expression on NK cells in the first trimester and found in comparison to normal pregnancies, patients with recurrent miscarriage had decreased Tim-3 expression and less anti-inflammatory cytokine production. In a mouse model, transfer of Tim-3 expressing NK cells reduced miscarriage rates. During the first trimester, Gal-9 levels increase and remain elevated throughout pregnancy. Preliminary data in a small population of patients who had miscarriages were found to have significantly lower Gal-9 levels detected at 8 weeks gestation compared to those who continued on to have a term pregnancy. Additionally there is evidence that Gal-9 increases production of interleukin (IL-4), and in patients with recurrent miscarriage IL-4 levels are decreased [5]. These identifications have prompted further investigation into predictive value of Gal-9 and IL-4 levels in early pregnancy on the outcome of a pregnancy.

Angiogenic factors, such as vascular endothelial growth factors (VEGF), have been shown to have a critical role in pregnancy at both the local and systemic level. Locally, CD56+ uterine natural killer cells within the endometrium express higher levels of VEGF in women with recurrent miscarriage versus women with proven fertility. Histological differences in VEGF expression and placental vascular bed patterns have been observed in tissue from patients with a miscarriage compared to women with a viable pregnancy. Serum VEGF levels are significantly different in patients with recurrent pregnancy loss than in women with proven fertility [8]. Serum VEGF levels, including VEGF-A, -C, and -D, are significantly higher at 8 weeks gestation in pregnancies that result in a spontaneous loss compared to pregnancies that go on to result in a live birth. Therefore, additional studies are needed to establish if early first trimester serum levels of VEGFs, Gal-9, and IL-4 are significantly different in pregnancies that result in miscarriage or live birth. Miscarriage affects approximately 15-20% of pregnancies. Therefore, establishing predictive factors of miscarriage could have implications for a large portion of couples and could serve to guide current and future family planning efforts. It is, essential to gain an understanding of mechanisms underlying infertility and miscarriage, so that more effective treatments and protocols can be developed.

The study hypothesis states low levels of Gal-9 and IL-4, and high levels of VEGF at the time of embryo transfer will be predictive of failed embryo transfer and spontaneous miscarriage. The primary endpoint is the maternal blood levels of galectin-9, IL-4 and VEGF-A,-C,-D on the day of embryo transfer in cycles that result in live birth versus no live birth, or that result in miscarriage compared to live birth.

Inclusion criteria

• Females, 18 years old or older
• Patients undergoing a frozen embryo transfer OR confirmed intrauterine pregnancy between 5 0/7 weeks and 9 6/7 weeks gestation
• Patients planning to have all bloodwork done at Mayo Clinic Rochester
• Patients planning to deliver at Mayo Clinic or within the Mayo Clinic Health Systems

Exclusion criteria

• Non English speaking
• Pregnancy with multiple fetuses
• Patient has non-viable pregnancy