Methods
This was a pseudo-longitudinal study (a variant of repeated cross-sectional study) conducted from January 2018 to January 2020 in two Ministry of Health-affiliated hospitals in Mexico City: a tertiary-level unit and a secondary-level unit. This study was approved by the pertinent Institutional Review Board (IRB; Register 2010/010/3117). Written informed consent was obtained from each patient prior to inclusion.
Women at different moments of spontaneous onset of the first stage of labor were included. Women with multiple pregnancy, chronic diseases, obstetric complications, and history of sexual intercourse or vaginal medication within the last 48 hours were excluded. Women with clinical or microbiological evidence of urinary tract infection or infectious vulvovaginitis, as well as those with clinical or laboratory evidence of premature rupture of membranes (PROM) and/or incomplete follow-up were eliminated.
Obstetrics and Gynecology specialists, previously standardized, evaluated the presence of clinical signs of spontaneous labor. Uterine activity was recorded with a tocodynamometer. Cervical changes and membrane integrity were assessed by vaginal examination. Gestational age was determined by the date of the last menstrual period. Clinical and demographic characteristics were obtained of all participants.
To represent initial changes associated to first phase of spontaneous labor, pregnant women between 12 and 41 weeks of gestation (WoG) were classified into one of the following five groups:
CVF samples were obtained by speculum examination. A Dacron tipped swab was placed into the posterior fornix and transferred to transporting buffer at 4ºC (0.05M Tris-base, 0.15M NaCl, 1% BSA, 0.1% Tween-20 and protease inhibitors). Samples were centrifuged at 2000 x g at 4ºC for 15 minutes and supernatant fluid was collected and stored at -80ºC.
Cytokines in the CVF were measured with the Milliplex Human Cytokine/Chemokine Magnetic Bead Panel and the MAGPIX Reader (Millipore, Burlington, MA, USA); including: (1) chemokines: CXCL8 (IL-8 or interleukin-8); (2) pro-inflammatory: interleukin-2 (IL-2), interleukin-12p70 (IL-12p70), interferon-gamma (IFN-γ), tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1β) and interleukin-6 (IL-6); (3) anti-inflammatory: interleukin-10 (IL-10), interleukin-4 (IL-4) and interleukin-1 receptor antagonist (IL-1RA).