P-366 Tumor necrosis factor-alpha and milk fat globule-epidermal growth factor 8: novel biomarkers to predict implantation failure and pregnancy loss
Authors: Alkon T , Yu L , Luna M , Hernandez-Nieto C , Sandler B , Asch R , Rusell H , Bocca S 
Published: June 2022

Abstract

Study question
Can implantation failure (IF) and pregnancy loss be predicted in serum prior to in vitro fertilization (IVF)?

Summary answer
Tumor necrosis factor (TNFα) and milk fat globule-epidermal growth factor 8 (MFG-E8) levels may serve as serum markers to predict IF and pregnancy loss.

What is known already
In a normal pregnancy, mediators such as TNFα are released creating a physiological inflammatory response. However, an exaggerated release of TNFα has been associated with IF and recurrent pregnancy loss (RPL). Recent studies demonstrated that TNFα up-regulates the expression of inflammatory factors such as MFG-E8. MFG-E8 is known to modulate implantation by acting at various levels of the trophoblast and endometrial compartments. Hence an overexpression of this protein may result in apoptosis, endometrial damage, and impaired implantation.

Study design, size, duration
This multicentric prospective controlled pilot clinical study was conducted from January 2016 to January 2020 and included 30 women in their natural cycle in which serum MFG-E8, TNFα, estradiol (E2), and progesterone (P4) levels were quantified in the early proliferative (cycle day 2) and secretory phases (urinary LH + 7 days). Additionally, an endometrial biopsy was obtained on urinary LH + 7 days for MFG-E8 and TNF α protein and gene expression analysis.

Participants/materials, setting, methods
Women ages 21-35y were recruited from 3 groups: fertile controls (C), unexplained IF (following 3 failed good quality embryo transfers), and RPL (at least 2 unexplained first trimester miscarriages). Patients with history of uterine surgery, abnormal uterine cavity (fibroids, endometrial polyps, adhesions, adenomyosis, and congenital uterine abnormalities), hydrosalpinx, diminished ovarian reserve, harboring chromosomal rearrangements, thrombophilia, or autoimmune diseases were excluded.

Main results and the role of chance
Ten women were included in each group. No statistical differences were found in age, BMI, AMH, baseline FSH, and baseline antral follicle count among cohorts. Mean serum E2 and P4 levels were similar among all groups in both the proliferative and secretory phases: E2 proliferative (C 69.19±26.64  pg/ml, IF 64.19±32.56  pg/ml, RPL proliferative 57.44±38.51; p = 0.55), E2 secretory (C 164.10±52.57  pg/ml, IF 172.57±121, RPL 173.81±.97.35; p = 0.25), P4 proliferative (C 0.45±0.15 ng/ml, IF 0.45±0.19 ng/ml, RPL 0.53±0.18 ng/ml; p = 0.85), P4 secretory (C 7.42±4.06 ng/ml, IF 7.8±4.56 ng/ml, RPL 8.05±4.38 ng/ml; p = 0.74). Mean serum TNFα levels were significantly higher in both, the proliferative and secretory phases for the RPL group (proliferative RPL 9.98±4.47  pg/ml, IF 4.73±2.56  pg/ml, C 3.42±1.01  pg/ml; p = 0.001 vs secretory RPL 8.67±4.45  pg/ml, C 3.35±0.94  pg/ml, IF 3.85±1.01  pg/ml; p = 0.03). Mean serum MFG-E8 levels were significantly higher in the IF group during the proliferative phase (IF 373±201  pg/ml, RPL 201±115  pg/ml, C 225.58±109.73pg/ml; p = 0.03), but not in the secretory phase (IF 237±101  pg/ml, RPL 189±116  pg/ml, C 199.41±112.43  pg/ml; p = 0.15). Endometrial MFG-E8 mRNA levels were significantly lower in the IF and RPL group compared to C (p = 0.03). TNFα mRNA levels were not statistically significant among groups (p = 0.12).

Limitations, reasons for caution
This is a pilot study to assess feasibility. Due to the small sample size, the effects of more subtle covariates would not have been detected. Future larger studies are warranted.

Wider implications of the findings
These novels differentially expressed serum and endometrial markers may provide information on the physiology of implantation and could generate the basis for non-invasive diagnostic tools and therapeutic use of MFG-E8/TNFα antagonists in women with IF and RPL.

Trial registration number
IIT-2014-100366