Immunomodulation During Pregnancy

The main objective of this study is to evaluate the effect of LDA on the modulation of innate immunity cells (NK cells, monocytes, γδ T cells) and/or acquired immunity (B and T lymphocytes, Treg cells, Th cells). The secondary objectives are to quantify pro-inflammatory and anti-inflammatory cytokines before and after LDA administration, as well as to assess maternal and fetal outcomes, comparing these parameters with those of low-risk PE pregnancies that do not require prophylactic LDA administration.

A prospective longitudinal study will be conducted, lasting a minimum of 40 weeks and a maximum of 24 months. Singleton pregnancies monitored at the Centro Hospitalar e Universitário de Coimbra will be included, with the first consultation occurring before 14 weeks of gestation and gestational age determined by first-trimester ultrasound. The study group will consist of women at high risk of PE according to clinical/historical criteria, who have not yet started LDA intake at the time of recruitment. Pregnant women will be classified as having a low or high risk of PE (risk of ≥ 1 in 50 for preterm PE) using the Fetal Medicine Foundation algorithm. This algorithm evaluates maternal clinical risk factors, mean arterial pressure (MAP), uterine artery pulsatility index (UtA-PI), and pregnancy-associated plasma protein A (PAPP-A). Women classified as high-risk will receive prophylactic aspirin (150 mg once daily).

As this is a preliminary study, 30 pregnant women will initially be included: 15 at high risk of PE (study group) and 15 at low risk (control group). Peripheral blood samples will be collected from both groups, utilizing routine pregnancy blood draws, at three time points (1st, 2nd, and 3rd trimesters). This will allow the evaluation of fluctuations in immune cells (NK cells, B and T lymphocytes, Treg cells, Th cells) and cytokine levels (IL-1, IL-6, IL-8, TNF-α, INF-γ, TGF-β, and IL-10). In high-risk PE pregnancies, additional peripheral blood samples will be collected before LDA administration and four weeks afterward.

Flow cytometry methodology will be used to determine immune cell frequencies, while plasma cytokine levels will be measured using ELISA and Luminex methodologies. After delivery, in both groups, histopathological evaluation (immunohistochemical staining) of the placenta will be performed, analyzing the aforementioned immune cells, as well as umbilical cord blood analysis using flow cytometry.

Inclusion criteria

- Singleton pregnancy

• Patients attending prenatal consultations at the Obstetrics Service A of CHUC, with the first consultation occurring before 14 weeks of gestation
• Gestational age determined by first-trimester ultrasound
• Study group: Women classified as high risk for PE according to clinical/historical criteria and the Fetal Medicine Foundation algorithm. LDA-naïve.

Exclusion criteria

Multiple pregnancy

• Autoimmune diseases
• Prior use of LDA or other immunomodulatory medication before potential recruitment
• History of spontaneous miscarriages and/or medical termination of pregnancy
• Fetal malformation